When sequencing DNA, what stops DNA synthesis in sequencing reactions?

When sequencing DNA, what stops DNA synthesis in sequencing reactions?

• A. Dideoxynucleotides
• B. DNA polymerase
• C. Insertion of thymine
• D. 3′-OH group

Answer: (A)

Dideoxynucleotides terminate DNA synthesis. These nucleotides lack the 3′-hydroxyl group, which is required for forming the next phosphodiester bond to extend the chain. When one is incorporated during sequencing, the polymerase cannot add further nucleotides, so the strand stops there. By including these terminators with normal nucleotides, a population of fragments ends at every possible base, and reading their lengths after separation reveals the sequence. The polymerase and normal bases would continue elongation if a 3′-OH were present, and termination is specifically due to the absence of that 3′-OH in the dideoxynucleotides, not because of thymine insertion.

Dideoxynucleotides terminate DNA synthesis. These nucleotides lack the 3′-hydroxyl group, which is required for forming the next phosphodiester bond to extend the chain. When one is incorporated during sequencing, the polymerase cannot add further nucleotides, so the strand stops there. By including these terminators with normal nucleotides, a population of fragments ends at every possible base, and reading their lengths after separation reveals the sequence. The polymerase and normal bases would continue elongation if a 3′-OH were present, and termination is specifically due to the absence of that 3′-OH in the dideoxynucleotides, not because of thymine insertion.