THE  RNA-­‐SEQ  ANALYSIS  PIPELINE  

Alicia  Oshlack  Murdoch  Childrens  Research  Ins5tute  

Genes  and  transcripts  

Gene  

transcript  

New genomic technologies are driving the science

Two  ways  to  look  at  sequencing  data  

Sequence  of(mapped)  read  • genome  sequencing    • variant  detec5on  • Muta5on  detec5on  • genomic  rearrangements  • Bisulfite-­‐seq  (methyla5on)  • RNA  edi5ng  etc.  

Posi5on  of  mapped  read  • RNA-­‐seq  • ChIP-­‐seq    • MeDIP-­‐seq  for  DNA  methyla5on  etc.  

   

4  

Two  ways  to  look  at  RNA-­‐seq  data  

Sequence  of(mapped)  read    • Assembly    • Determining  genes/transcripts    

Posi5on  of  mapped  read    • Expression  levels  • Differen5al  expression    

5  

Benefits  and  opportuni5es  of  RNA-­‐seq  

•  All  transcripts  are  sequenced  not  just  ones  for  which  probes  are  designed  (cf  microarrays)  

•  Annota5on  of  new  exons,  transcribed  regions,  genes  or  non-­‐coding  RNAs  

•  Whole  transcriptome  sequencing  –  The  ability  to  look  at  alterna5ve  splicing  –  Allele  specific  expression  –  RNA  edi5ng  

This  talk  

•  Analysis  of  RNA-­‐seq  data  for  the  purpose  of  determining  differen5al  expression  

•  How  much  are  expression  levels  changing  between  samples?  

RNA-­‐seq  

Pepke  et  al,  Nature  Methods,  2009  

Raw  data  

•  Short  sequence  reads    •  Quality  scores  

@SEQ_ID!GATTTGGGGTTCAAAGCAGTATCGATCAAATAGTAAATCCATTTGTTCAACTCACAGTTT!+!!''*((((***+))%%%++)(%%%%).1***-+*''))**55CCF>>>>>>CCCCCCC65  

RNA-­‐seq  analysis  steps  Raw  sequence  reads  

Map  onto  genome  

Summarize  reads  to  transcripts  

Sta5s5cal  tes5ng:  Determine  differen5ally  expressed  genes  

Systems  biology  

De  novo  assembly  Annota5on  based   Genome  guided  assembly  

Which  transcriptome?  

RNA-­‐seq  analysis  steps  Raw  sequence  reads  

Map  onto  genome  

Summarize  reads  to  transcripts  

Sta5s5cal  tes5ng:  Determine  differen5ally  expressed  genes  

Systems  biology  

De  novo  assembly  Annota5on  based   Genome  guided  assembly  

Which  transcriptome?  

Map  reads  to  the  genome  

•  Aligning  millions  of  short  sequencing  onto  the  3  billion  bases  of  the  human  genome  

•  Accuracy  vs  speed  •  Many  aligners  available  (BWA,  Bow5e,  STAR,  Novoalign,…)  

CDS   CDS   CDS   CDS  

CDS   CDS   CDS   CDS  

Gene  

transcript  

Sequencing  transcripts  not  the  genome  

Splice  site  mapping  •  Build  a  junc5on  library  from  all  combina5ons  of  known  exon  boundaries  

•  Determine  where  splice  junc5ons  occur  using  the  data  itself  -­‐  unbiased  by  annota5on.  

•  Several  so_ware  packages  to  do  this  such  as  TopHat,  SplitSeek,  SpliceMap…  

Gerber  et  al,  Nat  Methods,  2011  

Which  transcriptome  to  use?  

Raw  sequence  reads  

Map  onto  genome  

Summarize  reads  to  transcripts  

Sta5s5cal  tes5ng:  Determine  differen5ally  expressed  genes  

Systems  biology  

RNA-­‐seq  analysis  steps  

De  novo  assembly  Annota5on  based   Genome  guided  assembly  Which  transcriptome?  

Op5on  1  

•  Use  annota5on  (known  genes)  – Works  well  for  well  studied  organisms  (human,  mouse,  arabidopsis,  drosophila,…)    

– only  as  good  as  your  annota5on  – No  novel  transcripts  are  analysed  

Op5on  2:  Genome  guided  transcript  assembly  

•  Uses  the  loca5on  and  density  of  reads  along  the  genome  to  assemble  transcripts  

•  E.g.  Cufflinks  

•  Can’t  assemble  across  breaks  in  the  genome  – Cancer,  poor  genomes  

Op5on  3:  De  novo  transcriptome  assembly  

•  Assemble  transcripts  from  the  data  without  using  a  reference  genome  

•  “Harder”  than  genome  assembly  –  Orders  of  magnitude  varia5on  in  coverage  –  Con5gs  are  short  –  Alterna5ve  isoforms/transcripts  have  overlapping  sequences  –  *Very*  computa5onally  intensive  

•  So_ware  includes    –  Oases  (velvet)  –  TransAbyss  –  Trinity    –  …  

Example:  Annota5ng  the  chicken  W  chromosome  

   Z        Z      Z      W  

Male   Female  

Two  hypotheses  for  mechanisms  of  avian  sex  determina5on:  1.  Dominant  ovary  determining  gene  on  W  (cf  mammals)  2.  Dosage  of  Z-­‐linked  genes    

There  is  an  annotated  chicken  genome  

•  Chicken  W  chromosome  is  poorly  assembled  •  Are  genes  on  other  chromosomes  really  on  the  W,  in  par5cular  the  random  chromosome?  

Chromosome Assembled Size (Mb)

Size inc. random (Mb)

Estimated Size (Mb)

Estimated Genes

(Ensembl) Z 69 70 80 796

W 0.24 0.89 18-54 46

Un_random 56 - - 1287

                                                 Experimental  design      

   

PCR  Sexing  

+12hour  Blastoderms  

Hand  plate  for  PCR  Sexing  

RNA  

Stage  26  paired  gonads  (day  4.5)  

12  Female      

RNA  

16  Female  gonads  

Pooled  Samples  

12  Female  

16  Female  gonads  

12  Male  

12  Male  

16  Male  gonads  

16  Male  gonads  

RNA-­‐seq  • Illumina  HiSeq2000  • Paired-­‐end  100bp  • 4  lanes  • >80million  reads/sample  

 

Defining  the  transcriptome  

•  Annota5on  ~20,000  genes  •  Genome  guided  assembly  (Cufflinks)  ~45,000  genes  

•  De  novo  transcriptome  assembly  ~2.5  million  transcripts  (Abyss  with  filtering)!  

A  combined  approach  •  Assemble  cufflink  genes  using  transcripts  from  our  de  novo  assembly  

Annota5on  of  the  chicken  W  combined  all  three  approaches  

W/W_random ChromsomeUn_random ChromosomeAutosomes

Blastoderm CoverageGonads Coverage

Abyss TranscriptsCufflinks TranscriptsEnsembl Transcripts

1000 1500 2000 2500 3000 3500

base position

039

1

Coverage

Genome

EnsemblCufflinks

AbyssRASA1−W

1400 1600 1800 2000 2200

base position

012

6

Coverage

Genome

EnsemblCufflinks

AbyssST8SIA3−W

2000 2200 2400 2600 2800

base position

069

9

Coverage

Genome

EnsemblCufflinks

AbyssGOLPH3−W

0 500 1000 1500 2000 2500

base position

019

4

Coverage

Genome

EnsemblCufflinks

AbyssZSWIM6−W

0 200 400 600 800 1000 1200 1400

base position

082

Coverage

Genome

EnsemblCufflinks

AbyssNEDD4−like−W

Full  list  of  W  genes/transcripts  for  differen5al  expression  Ayers  et  al,  2013  

Summariza5on  

Take  your  “transcriptome”  and  add  up  the  reads  

Coding  Sequence   Exons   Introns   Splice  Junc5ons  

CDS   CDS   CDS   CDS  

§   Reads  in  exons  §   Exons  +  junc5ons  §   All  reads  start  to  end  of  transcript  §   De  novo  methods  

CDS   CDS   CDS   CDS  

CDS   CDS   CDS   CDS  

CDS   CDS   CDS  

Even  when  all  transcripts  are  “known”  summariza5on  or  expression  quan5fica5on  is  difficult.  How  do  you  assign  reads  to  transcripts?  

Transcript  1  

Transcript  2  

CDS   CDS   CDS   CDS  Transcript  3  

Summariza5on  turns  mapped  reads  into  a  table  of  counts  

Tag  ID! A1! A2! B1! B2!ENSG00000124208! 478! 619! 4830! 7165!ENSG00000182463! 27! 20! 48! 55!ENSG00000125835! 132! 200! 560! 408!ENSG00000125834! 42! 60! 131! 99!ENSG00000197818! 21! 29! 52! 44!ENSG00000125831! 0! 0! 0! 0!ENSG00000215443! 4! 4! 9! 7!ENSG00000222008! 30! 23! 0! 0!ENSG00000101444! 46! 63! 54! 53!ENSG00000101333! 2256! 2793! 2702! 2976!

…" …  tens  of  thousands  more  tags  …"

**  very  high  dimensional  data  **  

RNA-­‐seq  analysis  steps  Raw  sequence  reads  

Map  onto  genome  

Summarize  reads  to  transcripts  

Sta5s5cal  tes5ng:  Determine  differen5ally  expressed  genes  

Systems  biology  

De  novo  assembly  Annota5on  based   Genome  guided  assembly  

Which  transcriptome?  

Assessing  differen5al  expression  (DE)  

•  Which  genes  are  changing  in  their  abundance  between  samples?  

•  Sta5s5cal  tests  for  DE  (edgeR)  

Normaliza5on  

Accoun5ng  for/removing  technical  sources  of  varia5on  

Library  size  =  sum  of  each  column  

Tag  ID! A1! A2! B1! B2!ENSG00000124208! 478! 619! 4830! 7165!ENSG00000182463! 27! 20! 48! 55!ENSG00000125835! 132! 200! 560! 408!ENSG00000125834! 42! 60! 131! 99!ENSG00000197818! 21! 29! 52! 44!ENSG00000125831! 0! 0! 0! 0!ENSG00000215443! 4! 4! 9! 7!ENSG00000222008! 30! 23! 0! 0!ENSG00000101444! 46! 63! 54! 53!ENSG00000101333! 2256! 2793! 2702! 2976!

…" …  tens  of  thousands  more  tags  …"

Simple  thought  experiment  

•  Two  samples  A  and  B,  sequenced  to  the  same  depth  (same  library  size)  

•  Every  gene  that  is  expressed  in  A,  is  expressed  in  B  at  the  same  level  

•  Say  there  are  a  small  number  of  genes  that  are  expressed  uniquely  to  sample  B,  but  they  are  quite  highly  expressed  (lots  of  reads)  

•  Many  genes  within  the  common  set  will  appear  differen5ally  expressed  (B  <  A)  

Another  way  to  view  it  

•  Hypothe5cal  example:  Sequence  6  libraries  to  the  same  depth,  with  varying  levels  of  unique-­‐to-­‐sample  expression  

•  Differences  in  observed  counts  among  the  common  genes  

Red=low,  goldenyellow=high  

Marioni  et  al.  2008  RNA-­‐seq  data  

Distribu5on  of  log-­‐ra5os  of  counts    0  counts  are  omived    Red  line  =  trimmed  mean  

trimmed  mean  

•  5  lanes  of  liver  RNA,  5  lanes  of  kidney  RNA  •  Compare  two  single  kidney  libraries  (technical  replicates),  a_er  adjus5ng  for  library  size  

.

(a)

log2(Kidney1 NK1) − log2(Kidney2 NK2)D

ensi

ty

-6 -4 -2 0 2 4 6

0.0

0.4

0.8

log2(Liver NL) - log2(Kidney NK)

Den

sity

-6 -4 -2 0 2 4 6

0.0

0.2

0.4(b)

●

●

●

●

●

●

●

●

●

●

●●

●●

●●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●●

●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●●●

●

●

● ●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●●

●●●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●●●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●●

●●

●

●● ●

●

●

●

●

●

● ●

●

●●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●● ●

●

●●

●

●

●●●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●●●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●● ●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●● ●●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●●●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●● ●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●● ●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ● ●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●●

●

● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

● ●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●● ●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●●●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●● ●

●●

●

●

●

●●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●● ●●● ● ●

●

●●●

●●

●

●● ●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●●

●

● ●

●

●

● ●

●

●

●

●●

●

●

●● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●● ●

●

● ●●

●

●

●

● ●●

●

●

●

●● ●

●

●

●

●

●

●

● ●

●

●

●●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

● ●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●●

●

●

●

●

●

●●

●

●

●

●

●

●●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●

●

●

●●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●● ●

●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●●●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●● ●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●●

●

●●

●

●

●

● ●

●

●●

● ●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●●●●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

● ●

●

●

●

●

●●

●●

●

● ●●

●

●

● ●

●

●

●

●

●

●● ●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

● ● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●●

●

●

●

●

●

●

●●

●●● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●●●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●● ●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●●

●

● ●●

●

●

●

●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ● ●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

● ● ●●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

● ●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●● ●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●● ●●

●

●●

●

●

●

●

●●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●●

●

● ●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●●

●

● ● ●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●

● ●

●

●

●●

●

●●

●

●

● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

● ●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●● ●● ●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●●

●

●

●

●

● ● ●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

● ●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●●

● ●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

● ●●

●

●●●●●

●●●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

● ●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●●

●

● ●

●

●

● ●

●

●●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

● ●● ●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

● ●

●

●

●

●●

●

●

●●

●

●●

●

●

● ●

●

●

●

●

●

●●

●

●

●●

●

●

●

●●●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●●

●

●●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●● ●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●●●

●

●

●

●

●

● ●

●

● ● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●●

●

●

●

●

●

● ●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●●

●●

●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●●

●

●

●●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●●

●

●

● ●●

●

●●● ●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●●

●●

●

●

●

●

●

●

●

●

●●

●● ●●●

●●

●

●●

●●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●● ●

●

●

● ●●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●●●

●

●

●

● ●

●

●

●

●

●

●

●●

●● ●

●

●

● ●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

● ●

●

●

●

●

●

●

●

●● ●●

●

●●

●

● ●

●

●

● ●

●

●

●

●

●

● ●

●

●

●●●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●● ● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●●● ● ●●●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●●

●

●

●

●

●●●

●

●

●

●

● ● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●●

●

●

●

●●

●

●● ●

●●

●

●

●

●●

●

● ●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●●●

●

●

●

●

●

●

●

● ●●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●● ●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●

●

●

●

● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●●●

●

●●

● ●

●

●

●

●

●●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●

●

● ●

●

●

●●

●● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●● ●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●● ●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●● ●●● ●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●●

●

● ●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●●●

●

●

●

●

●●●●

●

●● ●

● ● ●

●● ●

●

●

●

●

●

●

●●

●

●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●●

●

●

● ●

● ●●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●● ●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ● ●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●●

●

●

● ●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●● ●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●● ●

●

●

●

●● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●●

● ●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●●

●●

●

●

●

●

● ●

●

●

●

●

●

●●●

●

●

●●

●

●

●●

●

●●

●●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

● ●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

● ●

●

●

● ●

●

●

●

●

●● ●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●●

● ●

●

●

● ●

●

●●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

● ●

●

●●

●

●

●

●●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●●

●●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●●

● ●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●●●

●●

●●●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

●●

●

●

● ●●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●●

●●

●●

●●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●●

●●

●

●

●●

●

●● ●

●

●●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

● ●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●● ●

●●

●

●

●

●

●

●●

●

● ●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●

●● ●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●●

●

●●

●

●●

●

● ●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

● ●

●

● ●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●● ●

●

● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●● ●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●●

●

●●

●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●● ●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

● ●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

● ●

●

●

●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ● ●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●● ●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

● ●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●●●

●

●●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●●

● ●

●

●

●

●●●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●●

● ●

●

●

●

●

●●

●

● ●●

●

● ●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●●

●

●●●

●

●●

●

●

●

●

●

● ●

●

●●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●● ●

●

●●●● ●

●

●

●

●

●

●●

● ●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●● ●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●● ●●

●

●●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●●

●●

●

●

●●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●●

● ●● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

● ●

●

●

●●

●

●●●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●●

●

● ●

●

● ●●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●●●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

● ●

●

●

●●●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

● ●●●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

● ●●

●

●

●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●● ●

●

●

●

● ●●

●

●

●

●

●

●

●

●●

●● ●● ●●

● ●

●

●●

●

● ●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●●

●●

●

●

● ●

● ●

●

●

●●●

●

●

●

●

●● ●

●

●●

●

● ●

●●

●

● ●

●

●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●● ●

●

●●

●

●●

●

●

●

●

● ●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

● ●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

● ●

●

●

●

● ●

●

●●

●●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

● ●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●●● ●

● ●

● ●

●●

●●

●

●

●

●

●●

●

●

●

●●

●●●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●●● ●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●●

●

●

●

●

●

●●

● ●

●

●● ●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●●

● ●

●

●

●

●●●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●●●

●●

●

●●

●

●

●●

●

●

●

●

●

● ● ●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●● ●

●

●

● ●

● ●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

● ●

●● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●●●

●●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

● ●

●

●

●

●

●●

●

●

●

● ●

●

● ●

●

●●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

● ●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

● ●●●●

●

●

●

●

●

●

●

●●●

●

●

●●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

● ● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●● ●

●

●●

●

●

●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●●

●

●

●

●

●

●●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●●●●

●●●

● ●●

●

● ●

●

●●

●

●

●● ●●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●

●●●

●

● ●

●

●

● ●

●●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●●

●

●

●

●●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

● ●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

● ●●

●

●●

● ●●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●● ●

●

●●

●

●

●

●●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●●

●

●

●●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●●●●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●●

●

●

●

●●●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●●

●

● ●●

●

●

●

●

●

●

●

●●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

● ●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●

● ●

●

●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●● ●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●● ●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ● ●

●

●

●

●

●

●

●

●●

●●●

●

●●

●

●

● ●●

●

●●

●

●

●●

●

●

●●

●

●

● ●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●●

●

●

● ●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

● ●

●●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●●

●

●● ● ●

●

● ●●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●●●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●●

●

●

●

●

●●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●

●

●●

●

●●●

●

●● ●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●●●

●

● ●

●

●●

●●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●●● ●●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

● ●● ● ●●

●

●

●

●

● ●

●

●

●●

●

●

●

●●

●

●

●

●●

●

●

●●●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●

●

● ●●

●

●

●●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●●●

●

●

●●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●●

●

●

● ●

●

●

●

●

●● ●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ● ●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

● ● ●●

●●

●

●

●

●

●●●

●●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ● ●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

● ●●

●

●

●

●

●

●

●●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

● ●● ●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

● ●●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

● ●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●● ● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

● ●

●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●●●

●●●

●

●

●●

●●●

●

●●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●●

●●

●●

●● ●

●

●

● ●

●●

●●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

● ●

●

● ●

●

●

● ●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●

●●

●

●

●

●

●

●●

●●

●●

●

●

● ●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●● ●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

-20 -15 -10

-50

5

A = log2( Liver NL Kidney NK)

M=

log 2

(Liv

erN

L)-

log 2

(Kid

ney

NK)

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

● ●

●

●●

●

●

●

●

●●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●●●

●● ●

●

●

●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

● ●

●●

● ●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●●

●

●●

●

●●

●●

●

●

●

●

●

● ●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●●

●

●

●

●

●

●

Housekeeping genesUnique to a sample

(c)

Same  sample.  Technical  replicates  

Different  samples.  Liver  vs  Kidney  

Another  way  to  view  the  data  

.

(a)

log2(Kidney1 NK1) − log2(Kidney2 NK2)

Den

sity

-6 -4 -2 0 2 4 6

0.0

0.4

0.8

log2(Liver NL) - log2(Kidney NK)

Den

sity

-6 -4 -2 0 2 4 6

0.0

0.2

0.4(b)

●

●

●

●

●

●

●

●

●

●

●●

●●

●●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●●

●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●●●

●

●

● ●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●●

●●●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●●●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●●

●●

●

●● ●

●

●

●

●

●

● ●

●

●●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●● ●

●

●●

●

●

●●●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●●●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●● ●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●● ●●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●●●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●● ●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●● ●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ● ●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●●

●

● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

● ●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●● ●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●●●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●● ●

●●

●

●

●

●●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●● ●●● ● ●

●

●●●

●●

●

●● ●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●●

●

● ●

●

●

● ●

●

●

●

●●

●

●

●● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●● ●

●

● ●●

●

●

●

● ●●

●

●

●

●● ●

●

●

●

●

●

●

● ●

●

●

●●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

● ●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●●

●

●

●

●

●

●●

●

●

●

●

●

●●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●

●

●

●●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●● ●

●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●●●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●● ●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●●

●

●●

●

●

●

● ●

●

●●

● ●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●●●●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

● ●

●

●

●

●

●●

●●

●

● ●●

●

●

● ●

●

●

●

●

●

●● ●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

● ● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●●

●

●

●

●

●

●

●●

●●● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●●●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●● ●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●●

●

● ●●

●

●

●

●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ● ●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

● ● ●●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

● ●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●● ●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●● ●●

●

●●

●

●

●

●

●●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●●

●

● ●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●●

●

● ● ●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●

● ●

●

●

●●

●

●●

●

●

● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

● ●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●● ●● ●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●●

●

●

●

●

● ● ●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

● ●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●●

● ●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

● ●●

●

●●●●●

●●●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

● ●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●●

●

● ●

●

●

● ●

●

●●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

● ●● ●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

● ●

●

●

●

●●

●

●

●●

●

●●

●

●

● ●

●

●

●

●

●

●●

●

●

●●

●

●

●

●●●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●●

●

●●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●● ●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●●●

●

●

●

●

●

● ●

●

● ● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●●

●

●

●

●

●

● ●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●●

●●

●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●●

●

●

●●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●●

●

●

● ●●

●

●●● ●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●●

●●

●

●

●

●

●

●

●

●

●●

●● ●●●

●●

●

●●

●●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●● ●

●

●

● ●●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●●●

●

●

●

● ●

●

●

●

●

●

●

●●

●● ●

●

●

● ●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

● ●

●

●

●

●

●

●

●

●● ●●

●

●●

●

● ●

●

●

● ●

●

●

●

●

●

● ●

●

●

●●●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●● ● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●●● ● ●●●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●●

●

●

●

●

●●●

●

●

●

●

● ● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●●

●

●

●

●●

●

●● ●

●●

●

●

●

●●

●

● ●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●●●

●

●

●

●

●

●

●

● ●●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●● ●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●

●

●

●

● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●●●

●

●●

● ●

●

●

●

●

●●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●

●

● ●

●

●

●●

●● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●● ●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●● ●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●● ●●● ●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●●

●

● ●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●●●

●

●

●

●

●●●●

●

●● ●

● ● ●

●● ●

●

●

●

●

●

●

●●

●

●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●●

●

●

● ●

● ●●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●● ●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ● ●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●●

●

●

● ●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●● ●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●● ●

●

●

●

●● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●●

● ●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●●

●●

●

●

●

●

● ●

●

●

●

●

●

●●●

●

●

●●

●

●

●●

●

●●

●●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

● ●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

● ●

●

●

● ●

●

●

●

●

●● ●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●●

● ●

●

●

● ●

●

●●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

● ●

●

●●

●

●

●

●●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●●

●●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●●

● ●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●●●

●●

●●●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

●●

●

●

● ●●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●●

●●

●●

●●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●●

●●

●

●

●●

●

●● ●

●

●●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

● ●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●● ●

●●

●

●

●

●

●

●●

●

● ●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●

●● ●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●●

●

●●

●

●●

●

● ●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

● ●

●

● ●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●● ●

●

● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●● ●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●●

●

●●

●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●● ●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

● ●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

● ●

●

●

●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ● ●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●● ●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

● ●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●●●

●

●●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●●

● ●

●

●

●

●●●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●●

● ●

●

●

●

●

●●

●

● ●●

●

● ●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●●

●

●●●

●

●●

●

●

●

●

●

● ●

●

●●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●● ●

●

●●●● ●

●

●

●

●

●

●●

● ●

●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●● ●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●● ●●

●

●●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●●

●●

●

●

●●

●●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●●

● ●● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

● ●

●

●

●●

●

●●●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●●

●

● ●

●

● ●●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●●●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

● ●

●

●

●●●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

● ●●●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

● ●●

●

●

●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●● ●

●

●

●

● ●●

●

●

●

●

●

●

●

●●

●● ●● ●●

● ●

●

●●

●

● ●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●●

●●

●

●

● ●

● ●

●

●

●●●

●

●

●

●

●● ●

●

●●

●

● ●

●●

●

● ●

●

●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●● ●

●

●●

●

●●

●

●

●

●

● ●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

● ●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

● ●

●

●

●

● ●

●

●●

●●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

● ●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●●● ●

● ●

● ●

●●

●●

●

●

●

●

●●

●

●

●

●●

●●●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●●● ●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●● ● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●●

●

●

●

●

●

●●

● ●

●

●● ●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●●

● ●

●

●

●

●●●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●●●

●●

●

●●

●

●

●●

●

●

●

●

●

● ● ●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●● ●

●

●

● ●

● ●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

● ●

●● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●●●

●●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

● ●

● ●

●

●

●

●

●●

●

●

●

● ●

●

● ●

●

●●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●●

●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

● ●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●●

●

●

●

●

●

●●

● ●●●●

●

●

●

●

●

●

●

●●●

●

●

●●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

● ● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●● ●

●

●●

●

●

●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●●

●

●

●

●

●

●●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●●

●

●●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●

●

●

●●●●

●●●

● ●●

●

● ●

●

●●

●

●

●● ●●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●●

●●●

●

● ●

●

●

● ●

●●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

● ●

●

●

●

●●

●

●●

●

●

●

●●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

● ●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

● ●●

●

●●

● ●●

●●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●● ●

●

●●

●

●

●

●●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●

●●

●

●

●●●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

●

●●

●●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●●●●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●●

●

●

●

●●●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●●

●

● ●●

●

●

●

●

●

●

●

●●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

● ●

● ●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●

● ●

●

●

●

●

●

●

●●

●

●●

●●

●

●

●

●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●●

●

●

●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●●

●● ●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●● ●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

●

●●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

● ●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ● ●

●

●

●

●

●

●

●

●●

●●●

●

●●

●

●

● ●●

●

●●

●

●

●●

●

●

●●

●

●

● ●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●●

●

●

● ●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●

● ●

●●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●●

●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●●

●

●● ● ●

●

● ●●

●

●

●

● ●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ●

●

●●●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●●

●

●

●

●

●●

●

●●

●

●

● ●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●●●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●

●

●●

●

●●●

●

●● ●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●● ●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

● ●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●●●

●●

●

●

●

●

●

●●●

●

● ●

●

●●

●●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●●●● ●●

●

●

●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

● ●● ● ●●

●

●

●

●

● ●

●

●

●●

●

●

●

●●

●

●

●

●●

●

●

●●●●

●

●

●

●

●●

●

●

●

●

●

●

●●

●

●

●● ●

●

●

●

●

●

●

●

●

● ●●

●

●

●●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●●●

●

●

●●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●●

●

●

● ●

●

●

●

●

●● ●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●

●●

●

●

●

●●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ● ●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

● ● ●●

●●

●

●

●

●

●●●

●●

● ●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ● ●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

● ●●

●

●

●

●

●

●

●●

●

●● ●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

● ●● ●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

● ●●

●

●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

● ●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

● ●● ● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

● ●

●

●

●

●●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

● ●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

● ●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●● ●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●●●

●●●

●

●

●●

●●●

●

●●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●

●

●

●●

● ●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

● ●

●

●●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●●

●●

●●

●● ●

●

●

● ●

●●

●●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●●

●

●●

●

●

●

●

●

● ●

●

● ●

●

●

● ●

●

●●

●

●●

●

●●

●

●

●

●

●

●

●

●●

●

●●

●

●

●●

●

●

●

●

●

●●

●●

●●

●

●

● ●

●

●

●

●

●

●●

●

● ●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●

●

●

●● ●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

● ● ●

●●

●

●

●

●

●

●

●

●

●

●

●

●

-20 -15 -10

-50

5

A = log2( Liver NL Kidney NK)

M=

log 2

(Liv

erN

L)-

log 2

(Kid

ney

NK)

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●●

● ●

●

●●

●

●

●

●

●●

●

●●●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●●

●

●●●

●● ●

●

●

●

●

●

●

●

●●

●●

●

●

● ●

●

●

●

●

●

●

●●●

●

●

●

●

●●

●

●

●

●

●●

●●

●

●

●

●●

● ●

●

●

●

●

●

●

●

●

●

●

●●

●●

●

●

●

●

●

●

●

●●

●

●

●

● ●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

● ●

●●

● ●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●

●●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●●

●

●

●

●

●

●

●

●●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●

● ●●

●

●

●

●

●●

●

●●

●

●●

●●

●

●

●

●

●

● ●

●

●

●●

●

●●

●

●

●

●

●●

●

●

●

●

●

●●

●

●

●●

●

●

●

●

●●

●

●●

●

●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

●

●

●

●

●

●●

●

●

●

●

● ●

●

●

●

●

●

●

●●

●

●

●●

●

●

●●

●

●

●

●

●

●

Housekeeping genesUnique to a sample

(c)

•  Sequencing  “real  estate”  is  fixed.  

•  Underlying  RNA  composi5on  can  be  very  different  –  e.g.  several  liver-­‐specific  genes  

•  An  adjustment  at  the  analysis  stage  should  be  made  

 

The  adjustment  to  data  analysis  is  a  scaling  constant  

•  Assump5on:  core  set  of  genes  that  do  not  change  in  expression.  

•  Pick  a  reference  sample,  compute  TMM  rela5ve  to  reference  •  TMM  (Trimmed  Mean  of  M  values)  M=log  ra5o    

•  Adjustment  to  sta5s5cal  analysis:  –  Use  addi5onal  offset  (GLM)  

Robinson  &  Oshlack,  Genome  Biology,  2010  

Sta5s5cal  tes5ng  for  DE  

•  For  EACH  GENE,  is  the  mean  expression  level  for  the  gene  under  one  condi5on  significantly  different  from  the  mean  expression  level  under  a  different  condi5on?  

Tag  ID! A1! A2! B1! B2!ENSG00000124208! 478! 619! 4830! 7165!ENSG00000182463! 27! 20! 48! 55!ENSG00000125835! 132! 200! 560! 408!ENSG00000125834! 42! 60! 131! 99!

…" …  tens  of  thousands  more  tags  …"

39  

DE  analysis  happens  in  R  •  Several  packages  currently  available  for  DE  analysis  of  RNA-­‐seq  data  in  R:  –  baySeq,    –  DEGSeq,    –  DESeq    –  edgeR  –  NBPSeq    (+  more  methods  with  less  refined  code,  e.g.  TSPM)  

•  edgeR  developed  at  WEHI,  so  this  is  my  favourite  •  edgeR  is  only  package  so  far  with  full  GLM  capabili5es  

•  Great  support  on  Bioconductor  mailing  list!  

41  

Counts  from  a  Single  RNA  Sample  Can  be  Modeled  with  Poisson  Distribu5on  

RNA  sample  

M  =  total  number  of  reads  ≈  20  million  (for  example)  λg    =  true  propor5on  of  gene  g  yg    =  number  of  reads  for  gene  g      

Read  1  Read  2  Read  3  Read  4  Read  5  Read  6  …  

Short  reads  

Map  reads  to  genome  

Davis  McCarthy  

42  

Counts  from  a  Single  RNA  Sample  Can  be  Modeled  with  Poisson  Distribu5on  

RNA  sample  

M  =  total  number  of  reads  ≈  20  million  λg    =  true  propor5on  of  gene  g  yg    =  number  of  reads  for  gene  g      

Large  M,  small  λi  à  yg  approximately  Poisson,  μg  =  Mλg  

Read  1  Read  2  Read  3  Read  4  Read  5  Read  6  …  

Short  reads  

Map  reads  to  genome  

What  we’re  interested  in  

What  we  observe  

Davis  McCarthy  

43  

A  Small  RNA-­‐Seq  Experiment  (Tech  Reps)  

Condi5on  A   Condi5on  B  

λg1   λg2   λg3   λg4  

yg1   yg2   yg3   yg4  

Genes  g  =  1,  …,  30k  

M1  

M2  M3   M4  

E(ygi)  =  Mi  λgi  Reads  Mi  ≈  20  million  

True  Technical  Reps  Show  Poisson  Varia5on  for  Each  Gene  

Data:    Marioni  et  al.,  Genome  Res,  2008   44  

Davis  McCarthy  

45  

A  Small  RNA-­‐Seq  Experiment  (Biological  Reps)  

Condi5on  A   Condi5on  B  

λg1   λg2   λg3   λg4  

yg1   yg2   yg3   yg4  

Genes  g  =  1,  …,  30k  

M1  

M2  M3   M4  

E(ygi)  =  Mi  λgi  Reads  Mi  ≈  20  million  

Davis  McCarthy  

Biological  Replicate  Data  shows  Quadra5c  Mean-­‐Variance  Rela5onship  

(development  cycle  of  slime  mould,  2  samples  at  hr00,  &  2  at  hr04)  

binned  variance,  sample  variance  

Data:  Parikh  et  al,  Genome  Biology,  2010  

46  Davis  McCarthy  

Need  to  account  for  biological  varia5on  

•  Technical  replicate  counts  for  a  gene  vary  according  to  a  Poisson  law,  i.e.  sequencing  varia5on  is  Poisson  

•  Biological  CV  (BCV)  is  the  coefficient  of  varia5on  with  which  the  (unknown)  true  abundance  of  the  gene  varies  between  RNA  samples.    

•  edgeR  models  a  quadra5c  mean-­‐variance  rela5onship  var(ygi)=  μgi  +  φgμgi

2  with  φg=Biological  CV2  

Robinson  et  al,  2010  

Output  of  analysis  

•  A  list  of  genes  with  p-­‐values  for  tes5ng  differen5al  expression  between  samples  

•  A  ranked  list  of  genes  

RNA-­‐seq  analysis  steps  Raw  sequence  reads  

Map  onto  genome  

Summarize  reads  to  transcripts  

Sta5s5cal  tes5ng:  Determine  differen5ally  expressed  genes  

Systems  biology  

De  novo  assembly  Annota5on  based   Genome  guided  assembly  

Which  transcriptome?  

RNA-­‐seq  analysis  steps  Raw  sequence  reads  

Map  onto  genome  

Summarize  reads  to  transcripts  

Sta5s5cal  tes5ng:  Determine  differen5ally  expressed  genes  

Systems  biology  

De  novo  assembly  Annota5on  based   Genome  guided  assembly  

Which  transcriptome?  

Learn  something!  

The  future  

•  Effec5vely  dealing  with  no  genome  or  poor  quality  genomes  (our  new  method,  Corset)  

•  Integra5ng  RNA-­‐seq  data  with  other  genomics  data  

•  Analysis  methodology  is  cri5cal  and  s5ll  developing  for  specific  purposes  

•  Opportuni5es  to  use  this  data  in  new  and  imagina5ve  ways  –  requires  new  analysis  methodology  

Acknowledgements  

•  Mark  Robinson  (Uni  Zurich)  •  Mavhew  Young  (Cambridge)  •  Nadia  Davidson  (MCRI)  •  Mavhew  Wakefield  (WEHI)  •  Gordon  Smyth  (WEHI)  •  Davis  McCarthy  (Oxford)