Fluorescent PCR detection method for simultaneously detecting deletion mutation and point mutation of genes by single tube

Abstract

The invention relates to the technical field of gene detection, in particular to a fluorescent PCR detection method for simultaneously detecting deletion mutation and point mutation of genes by a single tube. The method comprises the following steps: screening out available primers and probes, wherein the Tm value of the primers is higher than that of the probes; performing PCR reaction in the same reaction system of the single tube by using the obtained primers and probes; detecting change of the fluorescence value of the PCR reaction by using an instrument, analyzing and acquiring a channelfluorescence signal corresponding to the designed probe, and judging whether a detected sample has certain deletion mutation or not according to existence of melting peaks of all fluorescence channels; judging whether the detected sample has certain point mutation or not according to the temperatures of the melting peaks of all the fluorescence channels. The method has the beneficial effects thata plurality of deletion mutations and a plurality of point mutations can be simultaneously detected by only adding genome DNA of the detected sample into a detection reagent, one-time closed tube operation is realized, and pollution of a PCR product to an experimental environment is eliminated.

Description

technical field [0001] The invention relates to the technical field of gene detection, in particular to a fluorescent PCR detection method for simultaneously detecting gene deletion mutations and point mutations in a single tube. Background technique [0002] Single-gene genetic diseases refer to genetic diseases controlled by a pair of alleles. There are many kinds of them. More than 8,000 kinds have been found so far, and the combined incidence rate is as high as 1 / 100. Common single-gene genetic diseases include hereditary deafness, spinal muscular Atrophy, thalassemia, etc. Most single-gene genetic diseases can lead to death and disability, and there are very few effective drugs for treatment. Therefore, it is domestic recognized as the preferred preventive measure. Therefore, a simple, practical, accurate and sensitive gene mutation detection method is the premise and basis for the effective prevention and control of such genetic diseases. [0003] Common single-gene...

AI Technical Summary

Problems solved by technology

In general, gap-PCR and PCR-RDB, which are routinely used at present, must be detected separately, which is costly, heavy workload, cumbersome operation, low detection throughput, difficult to achieve automation and standardization, and there are defects after PCR amplification. The problem of laboratory carryover caused by open tube operation cannot meet the needs of large-scale population screening and clinical routine molecular diagnosis

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