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TT(N)mGCCTC inhibits archaeal family B DNA polymerases

Shuhui Sun1,2, Wei Guo1,2, Jin-Shu Yang1

  • 1College of Life Sciences, Zhejiang University, 866 Yuhangtang Road, Hangzhou, Zhejiang, 310058, P.R. China.

Scientific Reports
|February 2, 2018
PubMed

Insights

Certain DNA sequences, specifically TT(N)mGCCTC, can inhibit archaeal family B DNA polymerases. This finding is crucial for understanding polymerase chain reaction (PCR) fidelity and optimization.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Genetics

Background:

  • Archaeal family B DNA polymerases offer high-fidelity DNA replication crucial for techniques like PCR.
  • Thermostable proofreading DNA polymerases can sometimes fail in PCR amplification where Taq DNA polymerase succeeds.
  • Previous research indicated G-rich sequences forming G-quadruplex structures inhibit these proofreading polymerases.

Purpose of the Study:

  • To identify DNA sequences that inhibit archaeal family B DNA polymerases.
  • To investigate the mechanism of inhibition, particularly in single-stranded forms.
  • To differentiate inhibitory effects from Taq DNA polymerase.

Main Methods:

  • Investigated the inhibitory effects of specific oligonucleotide sequences on archaeal family B DNA polymerases.
  • Compared the inhibitory activity of TT(N)mGCCTC sequences against both archaeal family B DNA polymerases and Taq DNA polymerase.
  • Assessed the role of DNA secondary structures (single-stranded vs. G-quadruplex) in polymerase inhibition.

Main Results:

  • Identified single-stranded oligonucleotides with the sequence TT(N)mGCCTC that bind and inhibit archaeal family B DNA polymerases.
  • Demonstrated that Taq DNA polymerase is not inhibited by these TT(N)mGCCTC sequences.
  • Provided evidence that TT(N)mGCCTC inhibits thermostable DNA polymerases in a single-stranded form during PCR.

Conclusions:

  • The TT(N)mGCCTC DNA sequence is the first identified inhibitor of DNA polymerases in its single-stranded form.
  • Understanding these sequence-specific inhibitions can improve PCR protocols and enzyme selection.
  • This discovery offers new strategies for controlling DNA polymerase activity in molecular biology applications.

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