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Method and application for simultaneously detecting TCR (T cell receptor) and HLA (human leukocyte antigen) genotypes

A technique for genotyping, HLA-A

Pending Publication Date: 2021-12-03
深圳泛因医学有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0019] In addition, the conventional library construction scheme is to separate TRB and HLA to build a library, and only one of the results can be analyzed in the sequencing results. When HLA is needed to assist diagnosis, it is necessary to add a step of library construction, sequencing and analysis, which will cause Wasted time and increased costs

Method used

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  • Method and application for simultaneously detecting TCR (T cell receptor) and HLA (human leukocyte antigen) genotypes
  • Method and application for simultaneously detecting TCR (T cell receptor) and HLA (human leukocyte antigen) genotypes
  • Method and application for simultaneously detecting TCR (T cell receptor) and HLA (human leukocyte antigen) genotypes

Examples

Experimental program
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Embodiment 1

[0070] A method for simultaneous detection of TCR and HLA genotyping, the technical route process is as follows figure 1 shown, including the following steps:

[0071] 1. Sample collection

[0072] Use an EDTA tube to draw 5ml of peripheral blood, and turn the tube upside down several times to prevent coagulation.

[0073] 2. Sample extraction

[0074] Take 2ml of blood to the EP tube, use the blood DNA extraction kit (Hipure Blood DNAMidi Kit I, D312-02, Magen) and follow the instructions to obtain the DNA of the sample, use Q-bit to quantify it, and use it for DNA library construction.

[0075] 3. PCR amplification

[0076] Take 1.2 μg of the above DNA, add TCR amplification primers and HLA amplification primers at the same time according to the ratio in Table 1, and use multiplex PCR to capture and enrich the TCR VJ region and HLA-A, B, C 2, 3, 4 exon region.

[0077] In the previous work, the input amount of primers was adjusted and optimized in this step, and the dat...

Embodiment 2

[0115] A kit for simultaneously detecting TCR and HLA genotyping, comprising the TCR amplification primers and HLA amplification primers in Example 1.

Embodiment 3

[0117] A system for simultaneous detection of TCR and HLA genotyping, including:

[0118] A detection device, used for detection according to the method described in Example 1, to obtain sequencing data;

[0119] An analysis device, used to obtain the above-mentioned sequencing data for analysis and processing, and obtain TCR and HLA genotyping information;

[0120] The output device is used for outputting the TCR and HLA genotyping information.

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Abstract

The invention relates to a method and application for simultaneously detecting TCR (T cell receptor) and HLA (human leukocyte antigen) genotypes, and belongs to the technical field of gene detection. The method comprises the following steps: sample extraction: taking a biological sample to be detected, and extracting DNA in the biological sample; PCR amplification: taking the DNA, adding a TCR amplification primer and an HLA amplification primer at the same time, and amplifying a TCR target region and an HLA target region at the same time, wherein the HLA target region is a second exon region, a third exon region and a fourth exon region of the HLA gene; library construction: taking the amplification product to construct a library; sequencing on a machine: taking the library, and performing high-throughput sequencing on the machine; and data analysis: processing and analyzing the sequencing data to obtain TCR and HLA genotyping conditions. By adopting the method, the HLA-I type gene is amplified by the HLA primer group in the same PCR reaction system to perform typing identification under the condition of not increasing a library building step, and an HLA typing result is provided while TRB information is obtained, so that immune group library information is obtained to evaluate the health status of a sample provider.

Description

technical field [0001] The invention relates to the technical field of gene detection, in particular to a method and application for simultaneous detection of TCR and HLA genotyping. Background technique [0002] T Cell Receptor (TCR) is a heterodimer of molecular structure that T cells specifically recognize and bind antigen peptide-MHC molecules, and is composed of two different subunits. 95% to 99% of T cell receptors are composed of α subunit and β subunit, and 1% to 5% are composed of γ and δ subunits. This ratio will change with the development of the individual and the health status of the body. [0003] TRB (T cell receptor beta locus, T Cell Receptor Beta Locus) is a gene locus encoding the beta peptide chain in the TCR molecule. CDR3 of TRB consists of four gene regions, V (Variable), D (Diversity), J (Joining) and C (Constant), and each region contains several alleles. The gene rearrangement in the process of lymphocyte maturation forms various recombined sequen...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6806C12Q1/6869C12Q1/6883C12N15/11G16B20/30G16B30/00G16B50/30
CPCC12Q1/6806C12Q1/6869C12Q1/6883G16B20/30G16B30/00G16B50/30C12Q2600/16C12Q2537/143C12Q2531/113C12Q2535/122
Inventor 杨凡林莉娅张伟
Owner 深圳泛因医学有限公司
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