Laboratory of Molecular Genetics, KNU. Structure of the 4 bases found in DNA Base + sugar is called...
-
Upload
nigel-shelton -
Category
Documents
-
view
214 -
download
0
Transcript of Laboratory of Molecular Genetics, KNU. Structure of the 4 bases found in DNA Base + sugar is called...
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Structure of the 4 bases found in DNA
Base + sugar is called a nucleoside Base + sugar + phosphate is called a nucleotide
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Structure of A-T and G-C base pairsHydrogen-
bonds are shown as dotted red lines.A-T base
pairs have 2 and
G-C base pairs have 3
H-bonds
H-bonds are shown as thin
flat whitedisks in the
center
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
DNA strands are anti-parallel
The first proof was providedIn 1961 by measuring the
ratio of different dinucleotides in DNA. The concentration of
5’AG3’ was equal to 5’CT3’ (asexpected from an antiparallelorientation) and not equal to
5’TC3’ (as expected from a a parallel orientation).
DNA sequencing in 1970sconfirmed this conclusion.
5’
3’
3’
5’
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Structure of DNA
Watson and Crick in 1953 proposed
that DNA is a double helix in
which the 4 bases are base paired,
Adenine (A) with Thymine (T)
and Guanine (G) with Cytosine (C).
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
전기영동
+ -
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
DNA is negatively charged due to phosphates on its surface. As a result, it moves towards the positive pole.
Size separation of DNA fragments by electrophoresis in agarose gels
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
전기영동
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
전기영동 * Agarose gel 전기영동 시 DNA 의 이동에 영향을 주는 요인들 1) DNA 분자의 크기 느리게 이동한다 . 2) Agarose 의 농도 느리게 이동한다 . 3) DNA 형태 ( 구조 ) 에 따라 이동속도가 다르다 . supercoiled DNA, linear DNA, open circular DNA 의 순으로 빠르게 이동한다 . 4) 부하되는 전압이 이동속도가 빠르다 . 5) 전기장의 방향도 이동속도에 영향을 미친다 . 6) Ethidium bromide 는 DNA 이동속도를 15% 정도 감소시킨다 . 7) 전기영동 완충용액의 성분과 이온강도도 전기영동 속도에 영향을 준다 .
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Distance migrated by a DNA fragment in a gel is related to log10 of its size
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
pCDNA3.1 vector
Plasmid DNA analysis
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
PCR analysis
MCF7
MB231
MCF7
MB231
# 386
# 411
IgG
Inp
ut
H3K
9me3
H3A
c
SU
V42
0
H4K
20m
e3
DN
MT
1
KA
P-1
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
Siz
e M
arke
r 2 uL 4 uL 8 uL
SonicationRIPA lysis
- + - + - ++ + + + + +
Genomic DNA analysis
Laboratory of Molecular Genetics, KNULaboratory of Molecular Genetics, KNU
2) DNA 전기영동
Amount of agarose in gel
(%[w/v])
Efficient range of separation of linear DNA molecules (kb)
0.3 5~60
0.6 1~20
0.7 0.8~10
0.9 0.5~7
1.2 0.4~6
1.5 0.2~3
2.0 0.1~2
전기영동