Bioinformatics CourseDay 4

Perl Extensions:

BioPerland

Ensembl API

BioPerl

● Collection of Perl scripts and modules● Facilitate development of Perl scripts for

bioinformatics applications – not ready-to-use programs!

● Object-orientated Perl code● Generated by biologists,

bioinformaticians, computer scientists● Different levels of complexity

What are modules?

● Perl extensions● .pm ending● inserted via 'use' or 'require' statement● can be nested:

– Bio::DB::GenBank– /sw/lib/perl5/5.8.6/Bio/DB/GenBank.pm

● contain code and documentation● perldoc Bio::DB::GenBank

Applications for BioPerl

● Sequence retrieval and manipulation● Data re-formatting● Run and parse output from bioinformatics

programs:– Blast– ClustalW– Tcoffee– GenScan– HMMER– ...

What is an Object?

● Example: Bio::Seq Object

use Bio::Perl

$seqobj = get_sequence('swissprot', 'TLR4_HUMAN');

What is an Object?

● Example: Bio::Seq Object

use Bio::Perl

$seqobj = get_sequence('swissprot', 'TLR4_HUMAN');

load module new function available

generates object

What is an Object?

● Example: Bio::Seq Object

use Bio::Perl

$seqobj = get_sequence('swissprot', 'TLR4_HUMAN');

load module new function available

generates object

$sequence_part = $seqobj->subseq(1,100);

$translation = $seqobj->translate();

provides methods

What is an Object?

● Example: Bio::Seq Object

use Bio::Perl

$seqobj = get_sequence('swissprot', 'TLR4_HUMAN');

load module new function available

generates object

$sequence_part = $seqobj->subseq(1,100);

$translation = $seqobj->translate();

$trans_trunc_rev = $seqobj->trunc(100,200)->revcom->translate();

combinations possible

provides methods

BioPerl's Objects

● Sequence Objects (representation of various types of sequences):– Seq– PrimarySeq– LocatableSeq– RelSegment– LiveSeq– LargeSeq– RichSeq– SeqWithQuality

BioPerl's Objects

● Location Objects:– Where is a feature on a sequence?

● Alignment objects● Blast reports● ...

Objects vs Functions

● Different access method– Example: reading an EMBL file:

Function:

$seq = read_sequence($file, 'embl')

Object:

$seqio = Bio::SeqIO->new( -format => 'embl' , -file => $file );$seqobj = $seqio->next_seq();

Bio::Perl example

● Load the module

● Get the sequence

use Bio::Perl;

$seqobj = get_sequence('swissprot', 'TLR4_HUMAN');

● Now you have a Bio::Seq object!

Bio::Perl is not BioPerl!

● BioPerl:– whole collection of Bio-related Perl extension

● Bio::Perl– just one of many modules– “Easy first time access to BioPerl via functions”– “Functional access to BioPerl for people who

don't know objects”– limited functionality– nice starter

DB entry to Objects

● Conversion of parts of the data into objects:

ID TLR4_HUMAN STANDARD; PRT; 839 AA.AC O00206; Q9UK78; Q9UM57;

Bio::PrimarySeq

OS Homo sapiens (Human).OC Eukaryota; Metazoa; Chordata; Craniata; ...

Bio::Species

FT REPEAT 52 76 LRR 1..FT REPEAT 77 100 LRR 2..

Bio::SeqFeatureI

Bio::Seq – Formats

AB1,ABI ABI tracefile format ALF ALF tracefile format CTF CTF tracefile format EMBL EMBL format EXP Staden tagged experiment tracefile format Fasta FASTA format Fastq Fastq format GCG GCG format GenBank GenBank format PIR Protein Information Resource format PLN Staden plain tracefile format SCF SCF tracefile format ZTR ZTR tracefile format ace ACeDB sequence format game GAME XML format locuslink LocusLink annotation (LL_tmpl format only) phd phred output qual Quality values (get a sequence of quality scores) raw Raw format (one sequence per line, no ID) swiss Swissprot format

Access through Bio::Seq object

perldoc Bio::Seq (methods returning strings):

$seqobj->seq(); # string of sequence

$seqobj->subseq(5,10); # part of the sequence as a string

$seqobj->accession_number(); # when there, the accession number

$seqobj->alphabet(); # one of 'dna','rna',or 'protein'

$seqobj->seq_version() # when there, the version

$seqobj->keywords(); # when there, the Keywords line

$seqobj->length() # length

$seqobj->desc(); # description

$seqobj->display_id(); # the human readable id of the sequence

Derived Bio::Seq objects

perldoc Bio::Seq (methods returning strings):

$seqobj->trunc(5,10) # truncation from 5 to 10 as new object $seqobj->revcom # reverse complements sequence $seqobj->translate # translation of the sequence

● Example:

$seqobj = read_sequence($file);

if ($seqobj->alphabet eq 'dna' or $seqobj->alphabet eq 'rna') {

$revcom = $seqobj->revcom;write_sequence('', 'fasta', $revcom);

}

Other Bio::Perl features

● Remote Blast:$seqobj = read_sequence($file);

$blast_report = blast_sequence($seqobj);

write_blast(">blast.out", $blast_report);

● Also possible to run stand-alone Blast

Other Bio::Perl features

● Generate alignments:# load moduleuse Bio::Tools::Run::Alignment::Clustalw;

# define parameters@params = ('ktuple' => 2, 'matrix' => 'BLOSUM');

# build a clustalw alignment factory$factory = Bio::Tools::Run::Alignment::Clustalw->new(@params);

# Pass the factory a list of sequences to be aligned.$aln = $factory->align('TLRs.fa');

# $aln is a SimpleAlign object

Other Bio::Perl features

● Work with alignments:$aln->length$aln->no_residues$aln->is_flush$aln->no_sequences$aln->score$aln->percentage_identity$aln->consensus_string(75)

??????L?LS?N?I??????????L??L??L?L??N?????????????????F?????L??L?L??N???????????????L??L?L????????????????????????????????????????????????????????F??L??L??L?L????????????????L?????L???????????????????????????L?????????????????????L?????????????????????????????L????????????????????????????????????????

mostly Leucines conserved

Other Features

● sequence statistics (chemical description, residue count, word frequency)

● finding restriction enzyme sites● finding amino acid cleavage sites● show and add sequence annotation● gene detection● manipulate phylogenetic trees● statistics for population genetics

BioPerl Documentation

NAME Bio::Perl - Functional access to BioPerl for people who don't know objects

SYNOPSIS use Bio::Perl;

# will guess file format from extension $seq_object = read_sequence($filename);

# forces genbank format $seq_object = read_sequence($filename,'genbank');

# reads an array of sequences @seq_object_array = read_all_sequences($filename,'fasta');

# sequences are Bio::Seq objects, so the following methods work # for more info see Bio::Seq, or do 'perldoc Bio/Seq.pm'

print "Sequence name is ",$seq_object->display_id,"\n"; print "Sequence acc is ",$seq_object->accession_number,"\n"; print "First 5 bases is ",$seq_object->subseq(1,5),"\n";

More Info

● less `locate bptutorial` (or perl -d `locate bptutorial`)

● perldoc Bio::Perl

● www.bioperl.org

● BioPerl course: www.pasteur.fr/recherche/unites/sis/formation/bioperl/

Ensembl

● joint project between EMBL-EBI and the Sanger Centre

● automatic annotation on selected eukaryotic genomes

● free access to all the data and software● ww.ensembl.org

Ensembl Organisms

● Human● mouse● fly● worm● chicken● cow● rat

● dog● chimp● zebrafish● pufferfish● mosquito● honey bee● yeast

local installations (Arabidopsis)

Ensembl Website

Ensembl Website

Ensembl Pipeline

● Perl-based scripts ● run programs to detect and annotate

genes● compare genomes● provide Web graphics ● all data stored in MySQL database

Ensembl release cycle

● Data sets and software updates approximately ten times a year

● Versions for web code and databases ● All older versions (back to 2004)

accessible● Registry allows easy switch between

versions

Ensembl database

● very rich data set● complex database layout● user-friendly Web interface● DB components:

– Core– Compara

● abstract layers through API (Perl, Java)● anonymous @ ensembldb.ensembl.org

– e.g. homo_sapiens_core_38_36

Ensembl core

● Genome sequences and annotation info– Gene transcripts– Protein models

● Assembly information● CDNA and protein alignments● External references● Markers● Repeats regions

Other Ensembl data sets

● EST databases ● Variation databases● Both with application programming

interface (API), e.g. Perl modules

Ensembl compara

● Multi-species database● Genome-wide species comparison● Re-calculated for each release● Pair-wise whole genome alignments● Synteny sets● Orthologue predictions● Protein family clusters

Compara: Genome comparison

mysql> SELECT * FROM method_link;+----------------+----------------------+| method_link_id | type |+----------------+----------------------+| 1 | BLASTZ_NET || 2 | BLASTZ_NET_TIGHT || 3 | BLASTZ_RECIP_NET || 4 | PHUSION_BLASTN || 5 | PHUSION_BLASTN_TIGHT || 6 | TRANSLATED_BLAT || 7 | BLASTZ_GROUP || 8 | BLASTZ_GROUP_TIGHT || 101 | SYNTENY || 201 | ENSEMBL_ORTHOLOGUES || 202 | ENSEMBL_PARALOGUES || 301 | FAMILY |+----------------+----------------------+

● Which methods are available?

Compara: Genome comparison

mysql> SELECT * FROM genome_db WHERE genome_db_id IN (1, 11);+--------------+----------+-------------------------+------------+| genome_db_id | taxon_id | name | assembly |+--------------+----------+-------------------------+------------+| 1 | 9606 | Homo sapiens | NCBI34 || 11 | 9031 | Gallus gallus | WASHUC1 |+--------------+----------+-------------------------+------------+

● Which genomes are available?

mysql> SELECT * FROM method_link_species_set WHERE method_link_species_set_id = 71;+----------------------------+----------------+--------------+| method_link_species_set_id | method_link_id | genome_db_id |+----------------------------+----------------+--------------+| 71 | 1 | 1 || 71 | 1 | 11 |+----------------------------+----------------+--------------+

BLASTZ_NET (method_link_id = 1) has been used for linking all the species of this set: Human (genome_db_id = 1) and Chicken

(genome_db_id = 11).

● Which genomes were compared?

Compara: Genome comparison

More Info

● www.ensembl.org

● www.ensembl.org/info/software– Tutorials– Database schema outline

● Man pages, e.g.

perldoc Bio::Ensembl::DBSQL::DBAdaptor