Kit for CYP3A5 polymorphic site genotyping detection and detection method thereof

Abstract

The invention provides a kit for CYP3A5 polymorphic site genotyping detection and a detection method thereof. The kit comprises a wild type detection sequence 1, a wild type detection sequence 2, a mutant type detection sequence 1 and a mutant type detection sequence 2. According to the invention, a primer binding sequence and a probe binding sequence are integrated together, and the quenching effect is not ideal, so that double quenching groups are introduced in the design process of the sequences, the reaction sensitivity is improved, and the background signal noise is reduced; in order to improve the specificity of hybridization, on the basis of an original MGB probe, an rna basic group is introduced into the rear half region of the primer for distinguishing SNP typing detection sites, and mutation design is introduced into the upstream of the RNA modified basic group for further improving the specificity of the reaction.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a kit for genotype detection of CYP3A5 polymorphism sites and a detection method thereof. Background technique [0002] There are many methods for the detection of CYP3A5*3 gene polymorphism, such as restriction length polymorphism analysis (RFLP), direct sequencing method, blot hybridization method, Taqman technology, allele-specific amplification method (ARMS), etc. . Among them, the most commonly used sequencing method can directly detect the position and type of the mutation site, but this method has cumbersome operation steps, long detection cycle, low sensitivity, and the amplification product is prone to contamination. [0003] For example, in the probe hydrolysis method, two different fluorescently labeled probes are added to the PCR reaction system, and they can be completely paired with the two alleles respectively. Normally, fluorescence is quenched due to th...

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Problems solved by technology

[0006] The object of the present invention is to provide a method for CYP3A5 polymorphism in order to solve the problems of probe hydrolysis or ARMS-PCR, which have strict requirements on primer design region sequences and reaction enzymes, and are prone to non-specific amplification. The reagent kit and detection method for sex locus genotyping detection can improve the accuracy of detection results and the specificity of reaction

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