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Multiplex amplification system, and next generation sequencing typing kit and typing method for 21 micro-haplotype sites

A technology of multiple amplification system and typing method, which is applied in the direction of microbial determination/testing, sequence analysis, biochemical equipment and methods, etc., and can solve problems such as the loss of alleles of SNP sites and the inability to obtain typing results, etc.

Active Publication Date: 2019-09-10
HEBEI MEDICAL UNIVERSITY
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AI Technical Summary

Problems solved by technology

[0003] In view of the technical problems that the existing detection method STR typing technology cannot obtain ideal typing results, and the number of SNP sites may cause allele loss, the present invention provides a compound amplification method of 21 micro-haplotype sites. Increased system

Method used

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  • Multiplex amplification system, and next generation sequencing typing kit and typing method for 21 micro-haplotype sites
  • Multiplex amplification system, and next generation sequencing typing kit and typing method for 21 micro-haplotype sites

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Embodiment 1

[0053] The embodiment of the present invention provides a next-generation sequencing typing kit for 21 micro-haplotype sites, including next-generation sequencing custom panel kit, blood DNA extraction kit, Qubit dsDNA HS quantitative kit, next-generation sequencing Customized panel kits, real-time fluorescence quantitative kits, labcip quality inspection and analysis kits, and MiSeq on-machine sequencing reagents. The next-generation sequencing custom panel kit includes 53 single-end specific primers for 21 micro-haplotype sites (the nucleotide sequences are shown in SEQ ID NO.1~SEQ ID NO.53), and enzyme digestion reactions Buffer, FERA solution, enzyme mixture in enzyme cleavage reaction, ligation reaction buffer, ligation linker, DNA adapter, ligation solution, targeted PCR reaction buffer, and the 53 single-end specific primers for PCR amplification constitute a routine Forward and reverse primers for PCR, Taq DNA polymerase and conventional PCR reaction buffer.

Embodiment 2

[0055] The embodiment of the present invention provides a next-generation sequencing typing method for 21 micro-haplotype sites, which uses the next-generation sequencing typing kit for 21 micro-haplotype sites in Example 1 for sample detection . Including the following steps:

[0056] 1. Extract the DNA of the blood to be tested

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Abstract

The invention relates to the technical field of molecular biology, and especially relates to a multiplex amplification system, and a next generation sequencing typing kit and typing method for 21 micro-haplotype sites. The 21 micro-haplotype sites in the multiplex amplification system are derived from 21 autosomes, are balanced in allele frequency, are mutually independent between the sites, and have the advantages of polymorphism and low mutation rate. The kit contains 53 PCR-amplified single-end specific primers shown in SEQ ID NO. 1-SEQ ID NO.53, and can detect the 21 microhaplotype sites in a same reaction system. The multiplex amplification system, the kit and the typing method provided by the invention can solve the technical problem that a current STR typing technology cannot obtainan ideal typing result, and allele loss may occur due to excessive number of SNP sites.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a compound amplification system of 21 micro-haplotype sites, a next-generation sequencing typing kit and a typing method. Background technique [0002] Short tandem repeats (Short Tandem Repeats, STR) are the mainstream genetic markers for personal identification and kinship identification in forensic evidence laboratories. They have the characteristics of wide distribution, high polymorphism, and automatic typing. important role in judicial practice. Single Nucleotide Polymorphisms (Single Nucleotide Polymorphisms, SNP) is a sequence polymorphism caused by a single nucleotide variation in the genome. Compared with STR, it has a low mutation rate and short amplified fragments. It is more advantageous in material analysis. Moreover, SNP can also provide ancestral information, predict individual phenotypic characteristics, and provide more valuable clues for case detect...

Claims

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Application Information

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IPC IPC(8): C12Q1/6876G16B30/10
CPCC12Q1/6876G16B30/10
Inventor 李淑瑾丛斌周晶付丽红
Owner HEBEI MEDICAL UNIVERSITY
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