Kit for detecting RNA of hepatitis E virus

A technology of hepatitis E virus and detection kit, which is applied in the field of diagnosis of HEV infection in animals, and can solve problems such as short primers and low Tm values

Inactive Publication Date: 2009-09-23
BEIJING KINGHAWK PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the researchers did not report on the detection of different genotypes of HEV at present, and the primers they used in the SYBRGreen RT-PCR test were short (15-16mer), and their Tm values ​​were low, which may increase the potential non-genetic potential. Possibility of Target Fragment Amplification

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0114] Embodiment 1 kit test method

[0115] 1. Sample pretreatment

[0116] Serum and plasma samples were prepared according to conventional methods, and fresh samples were tested immediately, or frozen at -70°C to avoid repeated freezing and thawing.

[0117] Stool sample: Take 1.5-2.0 grams of stool into a 50ml centrifuge tube, add 30ml of PBS solution (containing 2% BSA), shake for 15 minutes, stand still for 2-3 minutes during this period, centrifuge at 3000rpm for 30 minutes, and take the suspension for testing or Freeze at -20°C.

[0118] 2. Nucleic acid extraction: (please operate strictly according to the requirements)

[0119] Note: a) Add absolute ethanol to the lotion A and lotion B bottles according to the label instructions before use, and mix well. b) Preheat the eluent at 70°C. c) Thaw the reagent reference substance and frozen samples at room temperature before use, shake and mix well. d) Differentiate the centrifugation settings (rpm and g) in the operat...

Embodiment 2

[0147] The preparation method of embodiment 2 kit (please give detailed preparation and packing process)

[0148] Preparation of kit components:

[0149] Nucleic acid adsorption column: purchased from outside, and used as a kit component after passing the quality inspection.

[0150] Cannula: 2.0ml nuclease-free cannula.

[0151] Precipitation aid: Outsourced and used as a kit component after passing the quality inspection.

[0152] Preparation of lysate: Take 590.8 g of guanidine isothiocyanate and 100 ml of Triton X-100, add them into 10 mM Tris / HCl (pH 8.0), dissolve and dilute to 1000 ml. Autoclave, after passing the quality inspection, sub-package as kit components.

[0153] Preparation of lotion A: Take 590.8 grams of guanidine isothiocyanate, add them into 10 mM Tris / HCl (pH 8.0), dissolve and dilute to 1000 ml. Autoclave, after passing the quality inspection, sub-package as kit components.

[0154] Preparation of lotion B: Weigh 1.57 g of Tris.base and dissolve it...

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PUM

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Abstract

The invention relates to a kit for detecting RNA of a hepatitis E virus (HEV). The kit can carry out qualitative detection to an HEV RNA in blood serum sample and a fecal sample of man and animals by adopting fluorescent PCR technology, wherein the HEV nucleic acid is separated by adopting extracted nucleic acid in a silicone gel membrane method, and the detection of the HEV RNA is realized by adopting the fluorescent PCR technology.

Description

Technical field: [0001] The invention relates to a kit and a detection method for in vitro qualitative detection of nucleic acid (RNA) of various genotypes of hepatitis E virus (HEV), which can assist the diagnosis and treatment of human clinical hepatitis E and the diagnosis of HEV infection in animals . Background technique [0002] Hepatitis E (HE, referred to as Hepatitis E) is similar to hepatitis A and is a self-limiting disease transmitted through the intestinal tract. It was not until 1980, when specific and sensitive hepatitis A antibody detection reagents were applied to the outbreak of epidemic hepatitis in water sources in India, that people realized that there was a virus that had similar epidemiological characteristics and clinical manifestations to HAV but was different from HAV. Known as enterally transmitted non-A non-B hepatitis virus. In 1983, Balayan, the former Soviet Union, first observed non-A, non-B hepatitis virus particles under an electron micros...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68G01N21/64C12R1/93
Inventor 曹健荣张誌闫宝山
Owner BEIJING KINGHAWK PHARMA
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