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SNP (Single Nucleotide Polymorphism) marker, primer and kit for evaluating solid organ transplantation condition and use method of SNP marker, primer and kit

A solid and state-of-the-art technology, applied in the field of SNP markers to evaluate the state of solid organ transplantation, can solve the problems of influence on the accuracy of results, low accuracy and sensitivity, and achieve reliable results

Pending Publication Date: 2022-04-12
成都仕康美生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] 3) If the puncture needle does not collect the lesion tissue part of the graft, it will have a great impact on the accuracy of the results, so the accuracy and sensitivity are not high

Method used

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  • SNP (Single Nucleotide Polymorphism) marker, primer and kit for evaluating solid organ transplantation condition and use method of SNP marker, primer and kit
  • SNP (Single Nucleotide Polymorphism) marker, primer and kit for evaluating solid organ transplantation condition and use method of SNP marker, primer and kit
  • SNP (Single Nucleotide Polymorphism) marker, primer and kit for evaluating solid organ transplantation condition and use method of SNP marker, primer and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0134] Using the kit provided by the invention to assess the risk of graft injury and rejection after solid organ transplantation

[0135] Take kidney transplantation testing as an example:

[0136] 1. Collect oral epithelial cells from kidney transplant recipients and donors respectively, extract DNA, and adjust the concentration to 10-40ng / ul

[0137] 2. Configure the SNP screening reaction system separately and perform PCR reaction; the reaction preparation is as follows:

[0138] Table 2

[0139]

[0140] 3. Add the configured reaction system to the 96-well plate for SNP screening as required, see figure 1 , (the 96-well plate includes: donor sample wells, recipient sample wells, negative control wells, and positive control wells arranged in sequence from left to right; donor sample wells, recipient sample wells, negative control wells, and positive control wells are all 3 columns and 8 rows)

[0141] 4. Then use the aluminum sealing film to cooperate with the seali...

Embodiment 2

[0167] Effect data:

[0168] Include 742 recipients after kidney transplantation, collect their plasma according to the method in Example 1, extract the total free DNA in the plasma, and perform digital PCR reaction. After the reaction, analyze each SNP site to obtain the percentage of GcfDNA, set 1 % is the reference value to distinguish graft injury or rejection from normal (graft injury or rejection (GcfDNA percentage > 1%) and graft normal group (GcfDNA percentage < 1%)).

[0169] Use MedCalc software to analyze, obtain the ROC curve ( figure 2 ), AUC=0.87, sensitivity=81.82%, specificity=79.59%.

[0170] The kit of the invention can timely, accurately and specifically reflect the health status of the kidney graft, and evaluate the graft injury and rejection risk after liver transplantation.

Embodiment 3

[0172] Include 416 recipients after liver transplantation, collect their plasma according to Example 1, extract the total free DNA in the plasma, and perform digital PCR reaction. After the reaction, analyze each SNP site to obtain the percentage of GcfDNA, set 10% Graft injury or rejection was distinguished from normal for reference values ​​(graft injury or rejection (GcfDNA percentage > 10%) and graft normal group (GcfDNA percentage < 10%)).

[0173]Effect data: use MedCalc software to analyze, obtain the ROC curve ( image 3 ), AUC=0.85, sensitivity=83.64%, specificity=71.43%.

[0174] The kit of the invention can timely, accurately and specifically monitor the health status of the liver graft, and evaluate the graft injury and rejection risk after liver transplantation.

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Abstract

The invention discloses an SNP (Single Nucleotide Polymorphism) marker, a primer and a kit for evaluating a solid organ transplantation condition and a use method of the SNP marker. The SNP marker consists of SNP loci which are homozygotes for a donor and a recipient and have different genotypes; according to the present invention, the specific SNP site of the transplanted receptor total free DNA is detected so as to directly obtain the content of the donor free DNA in the receptor free DNA, i.e., the GcfDNA percentage, and the transplantation rejection degree is rapidly and accurately determined according to the GcfDNA percentage, such that the kit can sensitively, specifically and accurately evaluate the solid organ transplantation condition, does not need the wound detection, and has the wide application range.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to SNP markers, primers, kits and usage methods for evaluating solid organ transplantation status. Background technique [0002] At present, the health monitoring of grafts after organ transplantation usually uses blood drawing for functional inspection, or tissue collection by puncture needle for pathological examination. [0003] For routine blood drawing function tests, the sensitivity and specificity of various indicators such as creatinine, ALT, AST, bilirubin, etc. are not high, and cannot accurately reflect the health status of the graft. [0004] For the current gold standard tissue biopsy, although it can directly reflect the health status of the graft. But there are the following major disadvantages: [0005] 1) Invasive detection, which causes great pain to the patient and damages the graft at the same time; [0006] 2) When the abnormality is detected, the substantial dam...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12Q1/6858C12N15/11
Inventor 魏亮
Owner 成都仕康美生物科技有限公司
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