Sanger chain-termination DNA sequencing requires deoxyribonucleoside triphosphates for synthesis. Which of the following are needed as the standard dNTP substrates?

Difficulty: Medium

Correct Answer: All of these (all four dNTPs are required)

Explanation:


Introduction / Context:
In Sanger sequencing, DNA polymerase extends a primer using normal deoxyribonucleotides (dNTPs) while occasional incorporation of a dideoxynucleotide (ddNTP) terminates extension. Understanding the role of both dNTPs and ddNTPs is essential.


Given Data / Assumptions:

  • The method is “chain-termination”.
  • Standard dNTPs drive processive synthesis; ddNTPs cause termination when incorporated.


Concept / Approach:
Polymerization requires the full set of dNTPs: dATP, dTTP, dGTP, and dCTP. ddNTPs are added at limiting ratios to terminate chains at specific bases. Without dNTPs, polymerase cannot extend, and sequencing would fail.


Step-by-Step Solution:

1) Identify the substrates for normal extension: all four dNTPs.2) Recognize ddNTPs are terminators, not the main substrates.3) Conclude that dATP, dTTP, dGTP, and dCTP are required.


Verification / Alternative check:
Standard protocols specify a mixture of all four dNTPs plus small amounts of labeled or unlabeled ddNTPs.


Why Other Options Are Wrong:

  • dATP only, or subsets like dGTP and dCTP, or dTTP only: incomplete; synthesis across diverse templates would stall at missing bases.
  • Only ddNTPs: would halt immediately; no readable ladders form.


Common Pitfalls:
Confusing the role of ddNTPs (terminators) with dNTPs (building blocks); overlooking that balanced dNTP pools are critical for uniform read lengths.


Final Answer:
All of these (all four dNTPs are required)

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