Constant-temperature nucleic acid hybridization solution based on RecA nucleoprotein filament, hybridization probe and application of probe

Abstract

The invention discloses a constant-temperature nucleic acid hybridization solution based on an RecA nucleoprotein filament, a hybridization probe and application of the probe. The hybridization probeis an RecA-nucleic acid complex and obtained through heterozygosis of RecA and a target nucleic acid fragment. The application method comprises the steps that the target nucleic acid fragment and theRecA are combined to form a mark nucleic acid probe; separation and purification of the mark nucleic acid probe are conducted; the mark nucleic acid probe is fixed; a hybridization reaction is conducted. According to the constant-temperature nucleic acid hybridization solution, by adopting a three-chain hybridization system, melting is not needed, and the complexity of reaction operation is lowered; constant-temperature hybridization is beneficial to design of an integrated instrument; the stability of a nucleic acid structure of the RecA-nucleic acid complex is improved, which is beneficial to preparation and storage of a detection chip; the matching accuracy and identification speed are high, and the hybridization reaction time is short; multi-hybridization of one locus and mass hybridization in a small range can be achieved; the mark diversity is achieved, and the detection flux is increased; the adaptability is high, and the application prospect is wide.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a constant temperature nucleic acid hybridization liquid based on RecA nucleoprotein filaments, hybridization probes and applications thereof. Background technique [0002] Nucleic acid hybridization technology is the most common analysis method in nucleic acid analysis. Its most common basic form is to use radioactive or non-radioactive labels to form probes with single-stranded DNA or RNA probes complementary to the target sequence. binding to the single-stranded target sequence, thereby proving the presence of the target sequence. [0003] The current nucleic acid hybridization technology first needs to melt the double-stranded nucleic acid. This operation requires a higher hybridization temperature, which increases the difficulty of the integration and automation of amplification and hybridization. [0004] Previous studies have found that after ATP-γ-S replaces ATP, the structu...

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Problems solved by technology

However, none of these studies used RecA as an independent hybridization probe for end-point hybridization

The patented technology with the application number 201710948491.6 uses RecA recombinase as the main substance in the reaction to react, but it uses RecA as a separate component for related hybridization reactions, adding a step of combining single-stranded nucleic acid with RecA, and the hybridization reaction is affected by RecA The influence of binding efficiency with nucleic acid increases the complexity of the hybridization step, and the labeling operation of the probe is more difficult

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