Hepatitis B virus multi-locus genotypic resistance mutation detection method

Abstract

The invention discloses a hepatitis B virus multi-locus genotypic resistance mutation detection method. A single tube nest type polymerase gene amplification method for detecting reverse transcriptase full gene sequence of virus includes that nest type PCR amplification is carried out in a single tube for twice, products of a first group PCR reaction are all used as templates of a second group PCR reaction by adjusting concentration of internal and external primers, and finally nest type PCR reaction is completed in the same single tube. The invention ensures specificity and economy of detection while obviously improving detection sensibility and has the detection range over a hundred million difference viral load.

Description

Technical field: [0001] The invention relates to a method for detecting drug-resistant mutations of hepatitis B virus multi-site genes, in particular to a single-tube nested polymerase chain reaction (PCR) amplification method for detecting hepatitis B virus (HBV) reverse transcriptase full gene sequence. Background technique: [0002] Nucleoside (acid) drugs are the main drugs for antiviral treatment of hepatitis B. Currently, the targets of anti-HBV nucleoside (acid) drugs in clinical use are all HBV reverse transcriptase (reverse transcriptase, RT), and long-term use will produce RT. For the problem of genetic drug resistance mutations, if drug resistance mutations are detected before medication, it can guide clinical rational drug use and avoid the impact of primary drug resistance mutations; if drug resistance is detected at the genetic level or early in virus breakthrough, clinical intervention can be carried out as soon as possible. Disease rebound can be avoided; ev...

AI Technical Summary

Problems solved by technology

The two rounds of traditional nested PCR reactions are carried out in two tubes respectively, resulting in the following disadvantages: first, it increases the chance of contamination of PCR amplification products; second, it is not suitable for simultaneous detection of samples with large differences in viral load; third, it increases testing cost

[0014] With the increase of the types of anti-HBV nucleoside (acid) drugs, the number and mutation forms of HBV drug-resistant mutations are also increasing, and the real-time fluorescent PCR method commonly used in clinical detection of YMDD mutations is difficult to detect multi-site gene mutations at the same time

[0015] However, there is currently no mature product for detecting HBV multi-site drug-resistant mutations on the market. Therefore, the development of PCR products with high sensitivity, wide detection range of viral load, strong specificity, simplicity, convenience, and low price that are easy for patients to accept directly Sequencing method to detect HBV drug resistance mutation is imminent

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