Kit and application thereof, and method and system for detecting area target variation

Abstract

The invention provides a kit, comprising a probe. The probe is fixed on a solid substrate or is free in a solution; the probe can specifically recognize a target area, wherein the target areas include one of the followings: at least one of the 163 genes shown in Table 1; or a CDS area of at least one gene in Table 1; or the upstream and downstream 10-20 bp area of a CDS area of at least one gene in Table 1. The invention also provides application of the kit, and a method and a system for detecting area target variation The kit and / or method and system can simply and conveniently obtain hereditary hearing impairment-related gene sequences at one time and high specificity, and accurately detect and analyze these related gene sequences, so that the detection and analysis results can assist the study of hereditary hearing impairment.

Description

technical field [0001] The present invention relates to the field of biomedicine, in particular, to a kit and its use, and more specifically, to a kit, the use of the kit, and a method and system for detecting variation in a target region. Background technique [0002] Hereditary deafness refers to deafness due to genetic and chromosomal abnormalities. This disease is deafness caused by changes in the genetic material (including chromosomes and genes located in it) of the parents and passed on to the offspring, and appears in a certain number of offspring. [0003] Currently, there are PCR-RFLP method, microarray chip method, ARMS-PCR method, fluorescence quantitative PCR method, gel electrophoresis method, fluorescence melting curve method, fluorescence capillary electrophoresis method for the detection of deafness gene. Microarray chip method and fluorescent quantitative PCR method are widely used at this stage, and their main technical defects are as follows: [0004] M...

AI Technical Summary

Problems solved by technology

[0004] Microarray chip detection requires whole blood DNA extraction (30min), PCR amplification (more than 2h), product hybridization analysis (more than 3h), the detection time is long (more than 6h), and the operation is cumbersome; due to too many operating procedures, It is easy to cause cross-contamination of samples; secondly, the recognition rate of a single base is relatively low, and false negatives and false positives are prone to occur, which affects the accuracy of the results; in addition, the cost of microarray chips is too high, requiring special chip scanning instruments, High detection cost; the amplification efficiency of ordinary ARMS-PCR products is only 1-10% of normal amplification, so the amplification system lacks stability; one reaction of four-primer ARMS-PCR can only detect one mutation site

The common disadvantages of common PCR products are: the genotype needs to be judged by the size of the band when interpreting the results, which lacks intuition

Gel electrophoresis detection results are prone to PCR cross-contamination, low detection sensitivity and time-consuming; using Taqman probes for SNP mutation detection requires 2 probes for each site detected, and has high requirements for probe specificity

The cost of probe synthesis is high, and there are requirements for the number of channels of the fluorescent PCR instrument, which is not conducive to popularization; the detection of one tube by the fluorescent melting curve method has few detection sites, and is limited by the channels of fluorescent dyes, which cannot meet the detection of multiple genes and multiple sites of deafness; PCR-RFLP detection technology is cumbersome to operate, requires enzyme digestion reaction, takes a long time to react, and requires gel electrophoresis detection, which is prone to cross-contamination of samples

Restriction sites are easily affected by gene variation; the multiplex PCR primer system is complex and difficult to design, and multiple pairs of primers will interfere with each other and competitively inhibit

Therefore, the more detection sites, the more primers required, the more difficult it is to optimize the PCR system. At the same time, due to the limitation of the fluorescence channel of the capillary electrophoresis instrument, it is difficult to promote the product. Generally, the maximum number of detection sites is no more than 20.

[0005] The common disadvantage of existing products is that there are few detection sites. They can only detect hotspot mutations in hereditary deafness genes, but cannot detect many rare sites. However, there are many deafness-causing genes and mutation sites. The above methods cannot meet the rapid technological development. , simple, wide range of detection requirements

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