Sequencing Technologies

Question 1

Dna Library Preparation

Answer 1

Dna is fragmented first
Adapeter ligated by DNA ligase to blunt ends

Question 2

How is target enriched to maximise coverage?

Answer 2

Pcr based methods-Illumina

Hybridisation- Whole EXOME sequencing

Question 3

How does sequencing by synthesis occur

Answer 3

By ILLUMINA

Free flow cell used to amplify DNA
Dna is primed by complementary sequence + DNA polymerase

4 individually labelled dNTP added to elongate strands and unbound dNTP removed

Forward/ reverse strands sequenced

To find 1 NTP responsible

Question 4

WGS

Answer 4

Identify mendelian + complex traits

Includes coding and non coding

Wider range of application

Question 5

WES

Answer 5

Exome is coding region contains 85% of DNA mutations

Highly penetrant mendelian disease

High coverage

Focuses on variation within exons and clinical diagnosis of genetic disorders

Question 6

Targeted NGS

Answer 6

Gene panels specific to certain disease types or cancers

Question 7

Prenatal testing

Answer 7

NIPT

Chromosomal abnormalities in cffDNA

Question 8

Pre implantation testing

Answer 8

PGT-A
Chromosomal abnormalities in embryonic biopsies

Question 9

Third generation sequencing

Answer 9

Pac bio
ONT- ultra long (nanopore)

long reads but less accuracy

No pcr so not affected by at and gc rich regions - prevents primers from binding together