II) NGS and Epigenomics

Question 1

What does epigenetic modifiés and what it doesn’t modify

Answer 1

Modify chromatin compaction and gene expression

Doesn’t modify DNA sequence

Question 2

3 divisions of sequencing

Answer 2

First generation
Second génération
Third generation

Question 3

Explain principle of BS ses

Answer 3

Bisulfite sequencing to id methylated cytosines (repression of transcription)

by converting them to uracil via deamynation using bisulfite treatment and comparison to sequences

Question 4

Principle of chIP seq

Answer 4

Chromatin immuniprecipitation sequencing to map global binding sites for a given DNA-binding protein or protein modification.

Antibody specific to TF or to histone modification after sample fragmentation.

Question 5

Principle of attack seq

Answer 5

Identification of accessible region in chromatin

Transposase in nucleosome free region. Fragmentation of chromatin and integration of adapters (tagmentation)
Only fragments with both types of adapters will be properly amplify and sequenced!

Question 6

What is the aim after all those epigenomic data’s collected ?

Answer 6

Integration of all together and coupling with RNA seq data for differential gene expression analysis and differential splicing of DNA.

Question 7

TAD ?

Answer 7

Topographically associating domains
Region with high interaction frequency

Question 8

CND ?

Answer 8

Chromatin nanodomain, where most enhancers promoter contact takes places

Question 9

Hi-C principle?

Answer 9

Method to study 3D genome organisation
Crossing dna, cut, bind with biotin, luggage, shear dna, sequence using paired ends.

Question 10

Sanger method ?

Answer 10

1st generation
Enzymatic synthesis and capillary electrophoresis to separated dna fragments.

Question 11

2nd NGS in a nutshell

Answer 11

Amplify of isolated DNA (no bacterial cloning)

Question 12

3rd NGS in a nutshell

Answer 12

Direct sequencing of single molecule template
Require less dna and no PCR

Question 13

Illumina ?

Answer 13

2nd generation, solid phase amplification (bridge amplification )

Détection of terminator
Colour émission
Cleave due and terminating group
repeat

Question 14

Two main types of 3rd generation sequencing

Answer 14

Single molecule real time sequencing
Nanopore sequencing

Question 15

Pacbio

Answer 15

Single molecule real time sequencing
Real time sequencing
Fluorescence pulse

Question 16

Two modes of SMRT

Answer 16

Continuous long reads
Circular consensus sequence

Question 17

Ore processing of reads

Answer 17

Base calling and Q score
FastQ format for reads