7. DNA sequencing

Question 1

Sanger sequencing

Answer 1

• add primer , nucleotide, 1 type of nucleotide, ddNTP, DNA polymerase
• make different length sequence because will be stopped when ddNTP is incorporated
• fragments will travel according to length

Question 2

Next generation sequencing

Answer 2

clonal display of DNA segments

Stepwise sequencing with optical detection

Question 3

HiSeq three steps

Answer 3

1. Library preparation
2. Cluster generation
3. Sequencing

Question 4

1. Library preparation

Answer 4

1. Fragment DNA
2. Repair ends, add A overhang
3. Ligate adapter
4. Select ligated DNA

Question 5

2. Cluster generation

Answer 5

1. Attach DNA to flow cell
2. Preform bridge amp
3. Generate cluster
4. Anneal sequence primer

Question 6

3. Sequencing

Answer 6

1. Extend first base
2. Read
3. De-block
4. Repeat to extend strand