midterm(review -...
TRANSCRIPT
Midterm ReviewBio5488 SP2017
Question 1 Total coverage:2x150 bp reads = 300 bp300 bp x 2e5 reads = 60e6 bp
Average coverage:60e6 bp / 12e6 bp = 5x
Poisson Distribution!
Accurate Call % (at least 4x)= 1 – {P(0,5) + P(1,5) + P(2,5) +P(3,5)}
Question 1 P(l) = 0.5
So, 5x in a row = 0.5 x 0.5 x 0.5 x 0.5 x 0.5 =0.0315
Question 5
ACmGTTCGCTTGAG
5’
3’
CTCAAGCGAACCmGT3’
5’
TGCmCAAGCGAACTC 5’3’
ACmGTTUGUTTGAG UTUAAGUGAAUCmGT
Bisulfite Treatment
Bisulfite Treatment
PCR/Sequencing PCR/Sequencing
ACGTTTGTTTGAG TTTAAGTGAATCGT5’ 3’
TGCAAACAAACTC3’ 5’ AAATTCACTTAGCA5’ 3’
3’ 5’
Homology
How many matches? Mismatches?
How many gap openings? Extensions? What are the penalties?
Alanine GCU, GCA, GCC, GCGTryptophan UGG
What is the likelihood that W-‐W occurs by chance in either?
What is the percent identity of the alignments?
Expression
Mixing up Type I/II error
Power analysis
• Increasing sequencing depth would not increase the statistical power
Epigenetics
• Q3• In general: DNAme ~ DNMT3b ChIP.• BUT, for lowly expressed genes, DNAme would not be that close to zero. • ALSO, DNAme in promoter region should be higher comparing to highly expressed groups
Epigenetics
• Q5:Interaction between Dnmt3b / methylated DNA and histone methyltransferases / H3K36me3 (recruitment of one by another); the associations are prior-‐differentiation-‐specific;DNA methylation in gene body prevents transcription from intragenic alternative TSS (see http://www.nature.com/nature/journal/v543/n7643/full/nature21373.html)
DNA binding specificity
• Strong binding to one nucleotide?• Frequency: 100%
• For any of the finger, background frequency is 0.25, while real frequency is 1. So: • Information content = 1.0*log2(1.0/0.25) = 2 bits• so 10 fingers are necessary.
• Why have specificity but degenerate?• Allow different level of affinity (diverse regulatory activity)• Allow easy on/off
Technologies used (and developed) to study genome folding
Adapted from http://web.uvic.ca/ail/equipment.htmlUnraveling the 3D genome: genomics tools for multiscale exploration. Viviana I. Risca and William J. Greenleaf. Trends in Genetics (2015)
Can be divided into two broad categories:
1. Imaging1. Bright-field2. Fluorescence3. EM4. Fluorescence in-situhybridization (FISH), etc.
2. Genome-wide1. DamID2. ChIA-PET3. Chromosome conformation capture-derived, etc.
HiC vs. ChIA-‐PET
• ChIA-‐PET: Chromatin Interaction Analysis by Paired-End Tag Sequencing
Nature Methods 6, 863 (2009)
?
Chromatin interaction
2D Graph of the same region