HBV detection method and kit, and application thereof

Abstract

The invention discloses an HBV detection method. The method comprises the following steps: separating a nucleic acid extract liquid from serum to obtain a sample to be detected; detecting a reaction result by a fluorescent quantitative PCR instrument, and setting a fluorescence signal as fluorescein corresponding to the 5' end fluorescence group of a Taqman fluorescent probe PgRNA-FP when collected, and collecting the fluorescence signal at 60 DEG C; and carrying out result analysis. Specific reverse transcription is carried out according to HBV PgRNA poly A, and then amplification is carried out according to HBV PgRNA specific primers and the probe, so possible interference caused by HBV DNA is effectively eliminated, and the HBV PgRNA detection specificity is effectively improved.

Description

technical field [0001] The present invention relates to a detection method, kit and application of bacteria, in particular to a detection method, kit and application of HBV virus. Background technique [0002] HBV virus is a double-stranded DNA virus. In the blood of hepatitis B patients, there are both HBV DNA and HBV RNA. kb, of which, the length of PgRNA is 3.5kb, and its sequence is highly homologous and overlapped with HBV DNA. It is very difficult for PCR specific selective amplification. To detect the content of PgRNA in blood (serum or plasma), the current conventional method is to extract The nucleic acid extracted from the serum of the hepatitis B patient was digested with DNaseI enzyme, and after the HBV DNA was digested, the remaining HBV PgRNA was subjected to PCR amplification to obtain the quantitative detection result of HBV PgRNA. However, for the digestion of high-concentration HBV DNA, it is necessary to repeatedly verify the dosage of DNaseI, so it is di...

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Problems solved by technology

However, for the digestion of high-concentration HBV DNA, it is necessary to repeatedly verify the dosage of DNaseI, so it is difficult to completely digest all concentrations of HBV DNA at one time, resulting in the remaining HBV DNA content interfering with the concentration of HBV PgRNA, resulting in inaccurate results If only the HBV PgRNA template can be amplified in a targeted and selective manner, the influence of HBV DNA on HBV PgRNA can be eliminated

[0003] In the prior art, HBV DNA and RNA co-extraction kits are usually used, and then the extracted nucleic acid is digested by DNaseI to remove HBV DNA and the remaining RNA. However, in reality, the concentration of HBV DNA varies greatly, and it is difficult to screen out a specific concentration. Elimination of all concentrations of HBV DNA, so it is difficult to remove high-load HBV DNA at one time, and it is necessary to repeatedly eliminate the impact of HBV DNA on HBV PgRNA. Such operations are time-consuming and laborious in clinical operations, and are likely to cause pollution. and quantitative inaccuracies caused by repeated digestion

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