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Kit for detecting genotypes of human platelet surface antigens HPA and HLA-AB

A HLA-A, genotyping technology, applied in the field of biomedical clinical molecular detection, can solve the problems of high cost, not suitable for wide application, etc., achieve high polymorphism, suitable for large-scale detection, and meet the effect of detection requirements

Active Publication Date: 2020-04-07
JIANGSU WEIHE BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It uses MGB probes and FAM / VIC to mark the a / b genotypes of HPA subtypes respectively, and judges the allele type according to the two fluorescence curves after amplification. This method is costly and requires a / b for each subtype Two MGB probes, not suitable for wide application

Method used

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  • Kit for detecting genotypes of human platelet surface antigens HPA and HLA-AB
  • Kit for detecting genotypes of human platelet surface antigens HPA and HLA-AB
  • Kit for detecting genotypes of human platelet surface antigens HPA and HLA-AB

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Experimental program
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Embodiment 1

[0055] 1. Raw materials and equipment

[0056] 1.1 Preparation of PCR reaction mixture: 0.18mM deoxynucleotide (dNTP), 1.8mM magnesium chloride (MgCl2), 60.3mM potassium chloride (KCl), 18.9mM Tris-hydrochloride (Tris-HCl) , glycerol (glycerol) 0.6% (v / v), dimethyl sulfoxide (DMSO) 5% (v / v) and formamide 2.5% (v / v), prepare 6*PCR reaction mixture 50mL according to the above ratio , Long-term storage at -20°C, temporary storage at 4°C for later use.

[0057] 1.2 Preparation of reaction plate:

[0058] 1.2.1 Preparation of primer-probe mixture: primer 0.54OD / mL, probe 1.05μM, make 6*primer-probe mixture.

[0059] 1.2.2 Spot the plate according to the table below: add internal standard primers and probes to each well, and the nucleotides are shown in SEQ ID No.197-199.

[0060]

[0061]

[0062]

[0063] The primer probe sequences and fluorescent labels of SEQ ID No.001-196 are shown in the following table:

[0064]

[0065]

[0066]

[0067]

[0068]

...

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Abstract

The invention provides a PCR amplification primer group and Taqman probe combination for detecting genotypes of HPA 1-6, 10, 15 and 21 and HLA-A and B, wherein the combination comprises 32 primer groups. The invention also provides a kit containing the combination of the PCR amplification primer group and the Taqman probe. The kit has the following technical effects: according to the invention, qualitative typing detection is carried out on HPA 1-6, 10, 15 and 21 genes and HLA-A and B gene loci by adopting an ARMS analysis method in combination with a Taqman multiplex fluorescence PCR technology; and the kit is improved on the basis of the traditional ARMS, overcomes various defects of the existing detection method, and has wide application prospect and clinical reference value.

Description

technical field [0001] The invention relates to a method and a kit for detecting human platelet surface antigen HPA and HLA-AB genotyping, belonging to the field of biomedical clinical molecular detection. Background technique [0002] Platelet is one of the formed components in mammalian blood, with irregular shape and no nucleus. Its main function is blood coagulation and hemostasis, and plays an important role in inflammatory response. There are a large number of antigens on the surface of platelets, which are usually divided into two categories: one is antigens shared with other blood cells or tissues, called platelet-associated antigens, such as HLA-I antigens, ABO antigen system, etc.; the other is platelet-specific antigens , showing the unique genetic polymorphism of platelets, that is, human platelet antigen (Human Platelet Antigen, HPA). [0003] HPA is an antigenic epitope located on the platelet membrane glycoprotein (glycoprotein, GP), and is an inherent antige...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6881C12Q1/6858C12N15/11
CPCC12Q1/6881C12Q1/6858C12Q2600/156C12Q2535/137C12Q2561/101C12Q2531/113
Inventor 陈炤源王浩张金涛章婷婷顾逸飞
Owner JIANGSU WEIHE BIOTECH
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