DNA Sequencing Quiz

DNA Synthesis and Sequencing Reaction

• The primer plays a crucial role in DNA synthesis during sequencing reaction by providing a starting point for DNA polymerase to synthesize new DNA strands.

dNTPs vs ddNTPs

• The main difference between dNTPs (deoxyribonucleoside triphosphates) and ddNTPs (dideoxyribonucleoside triphosphates) is that dNTPs have a 3' hydroxyl (-OH) group, while ddNTPs have a 3' hydrogen (H) instead, which prevents further DNA synthesis.

ddNTPs in DNA Sequencing Reaction

• The function of ddNTPs in DNA sequencing reaction is to terminate DNA synthesis by incorporating themselves into the growing DNA strand, making them ideal for Sanger sequencing.

Termination of DNA Synthesis

• Termination of DNA synthesis is base selective during sequencing reaction because each ddNTP is labeled with a different fluorescent dye, allowing for the identification of the incorporated nucleotide.

Incorporation of ddNTPs and dNTPs

• If a ddNTP is incorporated into the growing DNA strand during sequencing reaction, DNA synthesis terminates because the 3' end of the strand cannot be extended further.
• If a dNTP is incorporated into the growing DNA strand during sequencing reaction, DNA synthesis continues because the 3' hydroxyl (-OH) group allows for further extension.

Source of 3'OH Group

• The subsequent 3'OH group needed for DNA synthesis during sequencing reaction is provided by the incorporated dNTPs.

Role of Dyes Attached to ddNTPs

• The role of the dyes attached to ddNTPs during sequencing reaction is to label the terminating nucleotides, allowing for the identification of the incorporated nucleotide during Sanger sequencing.