Polymerase chain reaction detection method of hepatitis B virus genotyping

A genotyping and detection method technology, applied in the direction of microbial determination/inspection, biochemical equipment and methods, material stimulation analysis, etc., can solve the problems of product contamination, non-typing, time-consuming, etc., and achieve high coincidence rate and operation easy effect

Active Publication Date: 2011-08-03
THE FIRST AFFILIATED HOSPITAL OF FUJIAN MEDICAL UNIV
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Problems solved by technology

There are already some HBV genotyping methods, and the information provided by the sequencing method is comprehensive and reliable, but this method is cumbersome, time-consuming, and has high experimental conditions and requirements, so it is not suitable for widespread development
The results of other methods are not as reliable as the sequencing method, and they are still not standardized and commercialized. For example, PCR-RFLP is cumbersome to operate, and the restriction si

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  • Polymerase chain reaction detection method of hepatitis B virus genotyping
  • Polymerase chain reaction detection method of hepatitis B virus genotyping
  • Polymerase chain reaction detection method of hepatitis B virus genotyping

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Embodiment 1

[0023] 1. Object

[0024] In 2009, 186 inpatients and outpatients of the First Affiliated Hospital of Fujian Medical University, 124 males, with an average age of 35.21 years, and 69 females, with an average age of 35.71 years, were all positive for HBV DNA (10 3 ~10 8 copies / ml), stored at -80°C. All patients were excluded from hepatitis A, C and E virus infection and other diseases that cause liver lesions, and the diagnosis was in line with the diagnostic criteria revised by the Infectious Diseases and Parasitology Branch of the Chinese Medical Association (Xi'an) in 2000.

[0025] 2. Method

[0026] 1. Main experimental instruments and reagents: ABI7000 fluorescent quantitative PCR instrument (USA), viral genome DNA extraction kit (purchased from Shanghai Jierui Bioengineering Co., Ltd.), SYBR? Premix Ex Taq?? Engineering Co., Ltd.), Hepatitis B virus genotyping fluorescent PCR detection kit (purchased from Shanghai Fosun Medical Technology Development Co., Ltd.), and o...

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Abstract

The invention provides a polymerase chain reaction (PCR) detection method of hepatitis B virus (HBV) genotyping. The method comprises the following steps: a primer is designed according to the entire genome sequence of the HBV genotype; in the existence of the primer, DNAs are extracted from a serum sample to perform PCR amplification in a fluorescence PCR meter, the PCR product is analyzed according to the melting curve, the genotype is judged according to the Tm value of the PCR product and the HBV genotyping can be realized. The PCR melting curve method used in the invention is convenient to operate, the detection result of the method has higher coincidence rate with the detection result obtained by adopting a commercial genotyping kit; and the method can be used in the clinical detection of the HBV genotype. On the basis of the PCR technology, the PCR melting curve method which is to judge the HBV genotype according to the Tm value, is provided in the PCR detection method, thus the HBV genotype can be identified conveniently, rapidly and accurately.

Description

technical field [0001] The invention relates to a PCR melting curve detection method for HBV genotyping. Background technique [0002] Hepatitis B virus (HBV) mainly causes liver damage through the host's immune mechanism. The body's immune system recognizes viral antigens and attacks infected liver cells to cause inflammation. This process is affected by multiple factors including the host and the virus. Viral gene heterogeneity affects the expression of antigens and also plays an important role in this process. Different strains have different frequencies of certain mutations, different strains have different resistance to immune clearance of the body, and other factors may lead to different genotypes having different post-infection disease spectrum. [0003] HBV is usually divided into eight genotypes, A to H, based on the heterogeneity of the whole gene sequence ≥ 8%, or the heterogeneity of the S gene sequence ≥ 4%. HBV genotypes were distributed in certain geographic...

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68G01N21/64
Inventor 欧启水商红艳程祖建
Owner THE FIRST AFFILIATED HOSPITAL OF FUJIAN MEDICAL UNIV
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