Kit for detecting susceptibility gene of liver cancer of HBeAg negative HBV chronic infection liver cirrhosis patient and method

A chronic infection and kit technology, applied in the medical and health field, can solve the problem of no clinical prediction, no relationship between clinical outcomes, etc., and achieve monitoring that is conducive to prognostic goals, high sensitivity, and good specificity.

Active Publication Date: 2017-06-13
PEKING UNIV FIRST HOSPITAL +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the relationship between the rs187084 site of the TLR9 promoter sequence and the clinical outcome after HBV infection is not given, nor is it used to prompt clinical prediction after HBV infection.

Method used

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  • Kit for detecting susceptibility gene of liver cancer of HBeAg negative HBV chronic infection liver cirrhosis patient and method
  • Kit for detecting susceptibility gene of liver cancer of HBeAg negative HBV chronic infection liver cirrhosis patient and method
  • Kit for detecting susceptibility gene of liver cancer of HBeAg negative HBV chronic infection liver cirrhosis patient and method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1 Kit

[0048] 1. Amplification reaction solution

[0049] Used to amplify DNA containing SNP sites in samples, including: 10*buffer 0.625ul, Mg 2+ 0.325ul, dNTP0.25ul, Forward Primer lul, Reverse Primer 1ul, DNA 1ul (10ng-20ng), Hotstar taq enzyme 0.2ul.

[0050] Wherein, the Forward Primer (ie forward primer) is:

[0051] 5'-ACGTTGGATGTATTCCCCCTGCTGGAATGTC-3'.

[0052] Wherein, the Reverse Primer (ie reverse primer) is:

[0053] 5'-ACGTTGGATGTTACTATGTGCTGGGCACTG-3'.

[0054] Further, the reaction conditions for using the amplification reaction solution are: 95°C for 15 minutes; 45 cycles of 94°C for 20s, 56°C for 30s, 72°C for 60s, 72°C for 3min, and storage at 10°C.

[0055] 2. Purify the reaction solution

[0056] Used to purify the amplified DNA fragment, its main component is SAP enzyme, specifically: triple distilled water 1.53ul, Buffer0.17ul, SAP 0.3ul.

[0057] Further, the reaction conditions for using the purified reaction solution are: 37°C f...

Embodiment 2

[0062] Example 2 Method for Detecting Whether HBeAg-negative Patients Are Susceptible to Liver Cancer

[0063] 1. Specimen collection and DNA extraction

[0064] Take 200 μl of the subject's blood clot (the blood clot is dissolved in TES solution), and use the QIAGEN QIAampDNA BloodMini Kit to extract the whole genome DNA according to the operating instructions.

[0065] The specific operation steps are as follows:

[0066] 1) First add 200 μl of AL buffer into a 1.5ml centrifuge tube. Then the patient's specimen was gently blown and mixed, and 200 μl of the specimen was drawn into a centrifuge tube containing lysis buffer (AL buffer).

[0067] 2) Add 20 μl of QIAGEN protease or proteinase K to the centrifuge tube in step 1), shake and mix for 15 seconds, centrifuge at 8000 rpm / min for 10 seconds, and then put it into a 56°C electric thermostat water bath for 10 minutes.

[0068] 3) Centrifuge at low speed so that the liquid in the cap of the centrifuge tube enters the cent...

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Abstract

The invention provides a kit for detecting wherein an HBeAg negative HBV chronic infection liver cirrhosis patient is susceptible population to liver cancer or not and a method and relates to the field of biodetection. The kit comprises an amplification reaction liquid for amplifying a specific primer of DNA containing an SNP site in a sample, a purifying reaction liquid for purifying SAP enzyme of the DNA and an extending reaction liquid of a single basic group extending primer for obtaining a basic group detection result at rs187084 site of an TLR9 gene; when the detection result at rs187084 site of the TLR9 gene appears as a CT or / and TT genotype, the HBeAg negative HBV chronic infection liver cirrhosis patient is susceptible population to liver cancer. The specific primer and the extending primer provided by the invention can accurately amplify and extend a target gent, are high in sensitivity and good in specificity and can provide a ground for judging whether the HBeAg negative HBV chronic infection liver cirrhosis patient is developed to hepatocellular carcinoma or not, and the method is simple to operate, and a prognostic target is favorably monitored.

Description

technical field [0001] The invention relates to the medical and health field, in particular to a kit and method for detecting liver cancer susceptibility genes of HBeAg-negative HBV chronically infected liver cirrhosis patients. Background technique [0002] Currently, hepatitis B virus (HBV) infection is a worldwide public health problem. According to the statistics of the World Health Organization, there are 400 million HBV infected people in the world, and the HBV carriers in China account for 1 / 3 of the world. Among them, 10% of adults and about 80%-90% of children will develop chronic hepatitis B after infection, and chronic hepatitis B patients die of chronic hepatitis B complicated by liver cirrhosis and primary hepatocellular carcinoma. The phenomenon is determined by multiple factors and the interaction of various factors. Factors affecting the outcome of HBV infection include viral factors (such as viral genotype, viral load, and viral mutation); host immune statu...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6883C12Q2600/118C12Q2600/156
Inventor 曾争张独佾王彬彬王晶王熙
Owner PEKING UNIV FIRST HOSPITAL
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