New technique for preparing STR allelomorphic gene stairs and kit possessing the stairs

Abstract

The invention provides a new STR (short tandem repeat, Short tandern repeat,) preparation method of allele ladder (Allele ladder, AL), which comprises extracting DNA from human nucleated cells, and synthesizing corresponding to these STRs The primers of the site, select the DNA of individuals with different alleles, use PCR technology to specifically amplify the corresponding amplification products, use HPLC to purify and separate the amplified products, and perform PCR amplification→purification→PCR for multiple cycles Amplification→purification, and finally the purified products with different alleles at each site are mixed in equal proportions to form each STRAL. The present invention also provides a typing kit with the allele ladder prepared by this method. The preparation method and typing kit described in the invention can be used in aspects such as individual identification, paternity identification and gene diagnosis in the fields of forensic science, anthropology, genetics and diseases.

Description

technical field [0001] The present invention relates to forensic science, genetics, molecular biology, anthropology, clinical medicine, genomics, genetic diagnosis, genetic engineering and other technical fields, in particular to a preparation method of a new STR allele ladder and the A typing kit for allelic ladders. The technology can be used in the fields of forensic science, anthropology, genetics and oncology for individual identification, paternity test and genetic and tumor diagnosis. Background technique [0002] Human Genomic DNA has 3×10 9 bp, 10% of which are tandem repeats, known as satellite DNA. According to the length of the repeating unit, it can be divided into large satellites, medium satellites, small satellites and microsatellites. The repeat unit is a microsatellite composed of only 2-7bp, so it is also called short tandem repeat (short tandem repeat, STR). The number of satellite DNA repeat units in different human genomes is ...

AI Technical Summary

Problems solved by technology

[0005] (1) When the human genome is directly selected as the amplification template, due to the different lengths of the alleles, it is easy to have a dominant amplification phenomenon, that is, the alleles with shorter lengths are easy to be amplified, while the alleles with longer lengths are more likely to be amplified. The gene is not easy to be amplified, so that the amount of the amplified product is different, and the amount of each allele in the allelic ladder prepared from this is different, or even non-amplified, that is, no amplified product, which cannot be detected

[0006] (2) For mass production, it is necessary to frequently select and prepare templates that can represent all allele fragments. Not only is the workload large, but the purity and content of the same template used each time vary, making it amplified Conditions are difficult to standardize

[0007] (3) The molecular weight of human genomic DNA is large, the structure is extremely complex, and non-specific amplification is extremely prone to occur, and the product needs to be purified repeatedly, which is not suitable for mass production

[0008] (4) If human gene DNA amplification products are mixed and amplified again, the non-specific amplification will be more serious, and this method is not suitable for mass production

[0017] However, the above-mentioned short tandem repeat sequence allelic ladder preparation technology also has the following disadvantages: 1. Because of its many steps, it is complicated, and the preparation process takes a long time, so the probability of error is high; 2. Although the molecular cloning technology obtains The non-specific amplification products of the template are far less than those of human genomic DNA amplification products, but there are still interferences from non-specific DNA fragments (such as plasmid DNA), resulting in poor purity of the prepared allelic ladder

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