Epitaxial probe type qPCR detection method for SNV

Abstract

The invention discloses an epitaxial probe type qPCR detection method for single nucleotide variation (SNV). Four primers are designed according to known single nucleotide variation sites, two of the four primers are outer side primers, the other two primers are specific primers, bases at the 3' ends of the specific primers are respectively complementary with bases of a mutation template and a normal template, mismatch can be introduced or not introduced into bases near the 3' ends, epitaxy of a segment of artificial sequence is carried out from the 5' end of each specific primer, a homodromous affiliated primer is designed on the 5' segment of each artificial sequence, an affiliated probe is designed on the 3' segment of each artificial sequence, and in the amplification of the specific primers and downstream primers, the affiliated probes on the specific primers are degraded and emits light in the extension of the affiliated primers so as to indicate the amplification efficiency of the specific primers. If only one genotype of the single nucleotide variation sites is detected, the two outer side primers, one specific primer, the affiliated primer of the specific primer and the probes can be used. The method still has a plurality of implementation modes, and compared with probe type AS-PCR, the method has higher sensitivity and lower cost.

Description

technical field [0001] The invention relates to the field of gene detection, in particular to an extension probe type qPCR detection method for single base variation (SNV). Background technique [0002] Single nucleotide variation (Single nucleotide variants, SNV) is a single nucleotide variation in the DNA sequence. Single nucleotide variation widely exists in the human genome or other biological genomes, and it plays a regulatory role in biological processes such as RNA transcription and protein expression, and is closely related to human diseases. In this paper, SNVs include SNPs, which are not distinguished or given different names according to the population frequency of single base variation, whether the variation is from germ cells or somatic cells. [0003] There are many methods for SNV detection, such as PCR-Sanger sequencing, next-generation sequencing, pyrosequencing, TaqMan probe qPCR method, PCR-RFLP enzyme digestion method, Multiplex SNaPshot method, Sequenom...

AI Technical Summary

Problems solved by technology

The disadvantage of this variant is that the decrease in binding efficiency due to the introduction of deliberate mismatches on the specific primers cannot be compensated for by the amplification of the template provided by the two outer primers, resulting in most cases Reduced detection sensitivity and specificity

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