Double-stranded displacing probe high-specificity detection single-base change method based on thermodynamics optimization

Double-stranded displacing probe high-specificity detection single-base change method based on thermodynamics optimization

  • CN 105,132,528 A
  • Filed: 07/17/2015
  • Published: 12/09/2015
  • Est. Priority Date: 07/17/2015
  • Status: Active Application
First Claim
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1. the double-strand based on thermodynamic optimization replaces the method that nucleic acid probe high specific detects single sequence change, comprising:

  • (1) for the aim sequence of pre-detection, design double-stranded displacing probe, and design corresponding pcr amplification primer;

    Go out object target sequence by primer amplification, the reaction of the target sequence of probe and complete complementary is as reaction 1, and gained reaction product is product 1 and product 2;

    Probe with there is the reaction of target sequence of single sequence change as reaction 2, gained reaction product is product 3 and product 4;

    Calculate the standard Gibbs free energy Δ

    G of each nucleic acid molecule in reaction 1 and reaction 2 respectively 0, and the standard Gibbs free energy variation delta of reaction 1 and reaction 2 is calculated according to this rg 0;

    By probe and design of primers, make the free energy change amount of reaction 1 close to 0, reach theoretical optimum specificity;

    (2) according to the design of step (1), synthesize actual probe and primer, the high specific realizing object target sequence detects.

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