Question

Which sequencing technique relies on the synthesis of complementary strands in the presence of chain-terminating dideoxynucleotides?

• A. Sanger sequencing
• B. Next-generation sequencing
• C. Polymerase Chain Reaction (PCR)
• D. Pyrosequencing

Hint:

• Sanger Sequencing (Dideoxy / Chain-Termination Method)
  • Uses DNA polymerase, template, primer, dNTPs, and chain-terminating ddNTPs.
  • ddNTPs lack a 3'-OH group, so their incorporation stops DNA synthesis.
  • Generates a set of DNA fragments of different lengths, each ending with a specific ddNTP.
• This method was the dominant DNA sequencing technology for decades.

Answer 1

The Correct Option is A

Sanger sequencing is a method for determining the nucleotide sequence of DNA. This technique involves the synthesis of complementary DNA strands in the presence of chain-terminating dideoxynucleotides. Here's how it works:

1. Preparation: The DNA sample to be sequenced is denatured to produce single strands.
2. Primer Binding: A primer is annealed to the template strand, providing a starting point for the DNA polymerase.
3. Chain Extension: DNA polymerase extends the primer by adding complementary nucleotides.
4. Dideoxynucleotide Termination: The reaction mixture contains regular deoxynucleotides (dNTPs) and dideoxynucleotides (ddNTPs). When a ddNTP is incorporated, it prevents further extension, creating DNA fragments of varying lengths.
5. Separation and Analysis: The resulting fragments are separated by size using gel electrophoresis, allowing the sequence to be read based on the length of fragments ending with a specific ddNTP.

Sanger sequencing is the correct technique among the given options for synthesizing complementary strands with chain-termination.