Method for identifying genotype of insertion deletion locus of animal gene with HRM

Abstract

The invention discloses a method for identifying a genotype of an insertion deletion locus of an animal gene with HRM. The method comprises the following steps of searching potential gene insertion deletion locus information through an NCBI database, intercepting an upstream and a downstream of an insertion deletion fragment to design a specific primer, performing DNA mixed pool PCR amplificationsequencing, determining a real insertion deletion locus, performing no-other polymorphic site determination on the real existing insertion deletion locus, utilizing a high resolution melting curve (HRM) to perform individual genotype identification on an animal large group, and then performing PAGE verification and sequencing verification on an identification result through sampling. By adopting the method, the detection accuracy of gene insertion deletion is obviously improved, and the genotype identification efficiency is improved by about 3-5 times. Proved by a PCR and polyacrylamide gel electrophoresis method, typing performed by the method is reliable and effective. The method provides richer identification means to molecular marker detection work and provides a new reliable evaluation method to animal breeding work.

Description

technical field [0001] The invention belongs to the technical fields of molecular biology and cell engineering, and in particular relates to a HRM method for identifying the genotype of an insertion-deletion site of an animal gene. Background technique [0002] The genetic information of life is stored in the DNA sequence, and the entire DNA of each cell of a higher organism constitutes the genome of the organism. The variation of genomic DNA sequence is the basis of species genetic diversity and the original source of variation of biodiversity. There are a large number of insertion and deletion sites in the genome. Compared with the detection and application of single nucleotide polymorphisms, insertion and deletion are easier to be detected and have higher detection accuracy. They are regarded as a new molecular breeding marker and have The more obvious application value has attracted much attention. At present, there are certain limitations in the detection methods of g...

AI Technical Summary

Problems solved by technology

At present, there are certain limitations in the detection methods of gene indels. Direct sequencing and DNA fragment length polymorphism detection are mostly used. The former is expensive, while the latter is difficult to operate and takes a long time to identify. The positive probability is not conducive to the identification of gene insertion and deletion sites in large quantities

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