Chapter 5 Answers to self-check questions

5.1 What is the difference between a deoxyribonucleotide and a dideoxyribonucleotide and what is the function of each in the Sanger sequencing reaction?

A deoxyribonucleotide contains a hydroxyl group (OH) on position 3’ on the ribose sugar but lacks an oxygen on the second carbon hence why called a deoxyribonucleotide. A dideoxyribonucleotide instead will have only a hydrogen (H) on position 3’. It is therefore lacking two oxygens is therefore called a dideoxy.

5.2 What are the features that differentiate pyrosequencing from Sanger sequencing?

Pyrosequencing uses a combination of four different enzymes to produce a chemluminescence signal. Sequencing can be done in parallel allowing semi-high throughput. This technology does not need the use of ddNTPs tagged with fluorescent labels. Instead it uses ‘normal’ nucleotides.