Lecture 11: Next Gen Sequencing

Question 1

What are the pros and cons of sanger sequencing?

Answer 1

Pros:
- 500-800bp from a single read
- Highly reproducible
- High accuracy
- Relative simplicity
Cons:
- Lack of phase information
- Relatively expensive

Question 2

What are the 4 steps of NGS?

Answer 2

1. Fragment target DNA and ligate universal adaptors
2. Amplify single molecules
3. Sequence clonal amplicons
4. Assemble the data on the computer

Question 3

What are the characteristics of the Ion Torrent Sequencer?

Answer 3

• Post light: no fluorescence, no optics and no light
• Rapid
• Uses pH meter that detects changes in voltage

Question 4

What are the cons of Ion Torrent Sequencing?

Answer 4

• High cost per run
• Low sample number
• Has problems dealing with homopolymer and repeating regions