SNP mark applied to detecting RhD variation phenotype

A labeling and mutation technology, applied in the direction of recombinant DNA technology, DNA/RNA fragments, microbial determination/inspection, etc., can solve problems such as inability to judge, sample quality interference, easy missed detection, etc.

Active Publication Date: 2017-09-22
QINGDAO BLOOD CENT
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The second is that it is easy to miss the detection. Due to the variation of RhD antigen, the conventional antibody cannot react with it in the routine blood group serological test, so it is easy to be misjudged as RhD negative. Since there is no particularly effective measure to eliminate irregular antibodies, so This is very harmful to patients who need multiple blood transfusions and pregnant women
At present, the routine detection method of blood type is serological test, but this test is subject to many limitations, such as the quality of the sample, the interference of autoantibodies or irregular antibodies, the impact of diseases, etc. At the same time, for DEL type, it can only be tested by absorption and dissipation. The categories that can only be detected cannot be judged, which requires the use of molecular biology methods to diagnose at the gene level

Method used

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  • SNP mark applied to detecting RhD variation phenotype
  • SNP mark applied to detecting RhD variation phenotype
  • SNP mark applied to detecting RhD variation phenotype

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1: Screening of SNP markers

[0021] 1. Extraction of peripheral blood genomic DNA:

[0022] In compliance with the relevant national policies and regulations, and on the basis of the consent of the sampling subjects, draw 2-5mL of peripheral venous blood from RhD-negative or mutant blood donors, put them into EDTA anticoagulant tubes, and store them at -80°C for later use; frozen EDTA anticoagulant blood After the coagulated blood melts at room temperature, take 500 μL and put it in a centrifuge tube, add an equal volume of TE (pH 8.0), mix well, centrifuge at 10,000 rpm at 4°C for 10 minutes, and discard the supernatant.

[0023] Add 180 μL TE, 20 μL SDS (10%), and 8 μL proteinase K (10 mg / ml) to mix well, and place in a 37° C. water bath overnight. Remove the sample from the water bath and briefly centrifuge to pellet the sample. Add an equal volume of Tris-saturated phenol (about 300 μL) to the reaction tube, mix thoroughly, centrifuge at 10,000 rpm for 10...

Embodiment 2

[0037] Example 2: Genetic Validation of Parents of a Proband with a SNP Mutation

[0038] 1. Extraction of peripheral blood genomic DNA:

[0039] In compliance with the relevant national policies and regulations, and on the basis of the consent of the sampling subjects, draw 2-5mL of peripheral venous blood from RhD-negative or mutant blood donors, put them into EDTA anticoagulant tubes, and store them at -80°C for later use; frozen EDTA anticoagulant blood After the coagulated blood melts at room temperature, take 500 μL and put it in a centrifuge tube, add an equal volume of TE (pH 8.0), mix well, centrifuge at 10,000 rpm at 4°C for 10 minutes, and discard the supernatant.

[0040] Add 180 μL TE, 20 μL SDS (10%), and 8 μL proteinase K (10 mg / ml) to mix well, and place in a 37° C. water bath overnight. Remove the sample from the water bath and briefly centrifuge to pellet the sample. Add an equal volume of Tris-saturated phenol (about 300 μL) to the reaction tube, mix thoro...

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Abstract

The invention aims at providing an SNP mark applied to detecting an RhD variation phenotype. The SNP mark is that a 739th basic group from an initiation codon in an RHD gene encoding area mutates, G is mutated into C, a 247th amino acid is changed, and protein conformation is changed. According to the SNP mark disclosed by the invention, an SNP locus is obtained through detecting and screening, so that an effective path to perform RhD blood type gene diagnosis is provided; an application effect shows that an SNP locus and a detection primer of a gene disclosed by the invention can be effectively applied to clinical patients and blood station blood donators to quickly detect RHD genes.

Description

technical field [0001] The invention belongs to the technical field of gene diagnosis products, and in particular relates to a SNP marker for detecting red blood cell RhD variant blood type. Background of the invention [0002] The Rh blood group system (Rhesus monkeys) was first discovered by the famous scientist and Nobel laureate Karl Landsteiner from the red blood cells of rhesus monkeys, hence the name. The Rh blood group system is the most complex among the 36 human blood group systems, its importance is second only to the ABO system, and it plays an important role in clinical blood transfusion and diagnosis and treatment of hemolytic disease of the newborn. The D antigen in the Rh blood group system is the most immunogenic and the most complex polymorphism. For the Han people in mainland China, the RhD-negative population only accounts for about 3 / 1000. Because it is very rare, it is also called panda blood. Due to the strong antigenicity of RhD, RhD-negative people...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6881C12Q2600/156
Inventor 胡彬
Owner QINGDAO BLOOD CENT
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