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JC virus detection method, kit and application of kit

A detection method, JC virus technology, applied in the direction of determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve problems such as inability to achieve, and achieve the effects of preventing pollution, high sensitivity and low cost

Inactive Publication Date: 2016-10-26
BEIJING SEARCH BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0014] The main problem that this application solves is to provide a kind of detection method and test kit of JC virus, to solve the technical problem that cannot realize

Method used

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  • JC virus detection method, kit and application of kit
  • JC virus detection method, kit and application of kit
  • JC virus detection method, kit and application of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] This embodiment provides the method for obtaining the sample to be tested:

[0055] JCV DNA was extracted from human urine samples by magnetic bead method to obtain samples to be tested. The magnetic bead method extracts nucleic acid using a unique lysate and proteinase K to quickly lyse the cells and inactivate intracellular nucleases, then the genomic DNA is selectively adsorbed to the magnetic beads, and then through a series of rapid rinsing-separation steps, the inhibitor removal solution and washing solution to remove impurities such as cell metabolites and proteins, and finally the pure genomic DNA can be eluted from the magnetic beads with double distilled water.

[0056] 1. Reagents and components used to extract JCV DNA from human urine samples by magnetic bead method

[0057] (1) Lysis solution: 100mL dosage as an example, 40.47g of guanidine hydrochloride, 0.5132g of tris(hydroxymethyl)aminomethane, 0.0932g of sodium chloride, 6.78mL of Tween-20, add 16mL o...

Embodiment 2

[0077] Qualitative detection of JC virus

[0078] Present embodiment 2 provides a kind of qualitative detection method of JC virus:

[0079] 1. PCR amplification reaction of JCV DNA

[0080] 1) Reagents and components used in the PCR amplification reaction of JCV DNA

[0081] (1) JCV primer probe mixture:

[0082] The specific composition of the JCV primer-probe mixture is: upstream primer: JCV-F (50 μM) 0.6 μL, downstream primer: JCV-R (50 μM) 0.6 μL, Taqman fluorescent probe JCV-FP (20 μM) 0.5 μL, purified water 3.3 μL.

[0083] The preferred components of the JCV primer-probe mixture are: upstream primer JCV-F (50 μM) 0.6 μL, downstream primer JCV-R (50 μM) 0.6 μL, Taqman fluorescent probe JCV-FP (20 μM) 0.5 μL, internal standard upstream Primer (20 μM) 0.5 μL, internal standard downstream primer (20 μM) 0.5 μL, internal standard probe (10 μM) 0.375 μL, purified water 1.925 μL.

[0084] The upstream primer JCV-F comprises the base sequence shown in SEQ ID NO:1;

[008...

Embodiment 3

[0108] Present embodiment 2 provides a kind of quantitative detection method of JC virus:

[0109] 1. PCR amplification reaction of JCV DNA

[0110] 1) Reagents and components used in the PCR amplification reaction of JCV DNA

[0111] (1) JCV primer probe mixture:

[0112] The components of the JCV primer-probe mixture are: upstream primer: JCV-F (50 μM) 0.6 μL, downstream primer: JCV-R (50 μM) 0.6 μL, Taqman fluorescent probe JCV-FP (20 μM) 0.5 μL, internal standard Upstream primer (20 μM) 0.5 μL, internal standard downstream primer (20 μM) 0.5 μL, internal standard probe (10 μM) 0.375 μL, purified water 1.925 μL. The said upstream primer JCV-F comprises the base sequence shown in SEQ ID NO:1;

[0113] The downstream primer JCV-R comprises the base sequence shown in SEQ ID NO:2;

[0114] The Taqman fluorescent probe JCV-FP comprises the base sequence shown in SEQ ID NO: 3, the fluorescent reporter group is 6-FAM; the fluorescent quencher group is BHQ1.

[0115] (2) PCR r...

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Abstract

The invention discloses a JC virus detection method. The JC virus detection method comprises the following steps: extracting JCV DNA from human urine sample by utilizing a paramagnetic particle method, thereby obtaining a sample to be detected; performing PCR amplification reaction; utilizing a fluorescent quantitative PCR meter to detect reaction results, and when collecting fluorescent signals, setting fluorescein corresponding to a fluorescent group at Taqman fluorescent probe JCV-FP 5' end to collect the fluorescent signals at 60 DEG. The invention further provides a kit for detecting the JC virus. The detection method is high in sensitivity, can detect to 500 copies / mL at the lowest extent; and meanwhile, the specificity of the detection method is good, and according to the detection method, and no cross reaction with other virus occurs, such as BK virus (BKV), hepatitis C virus (HCV), human cytomegalovirus (HCMV), and hepatitis B virus (HBV).

Description

technical field [0001] The present invention relates to a virus detection method, a kit and its application, in particular to a JC virus detection method, a kit and its application. Background technique [0002] The JC virus belongs to the human polyomavirus branch of the polyomavirus species of the Papovaviridae family. It is very similar to the size structure and DNA sequence of the human polyomavirus SV40 (rhesus virus) and JC virus genome. It has no envelope and a coat Shell protein, containing a circular small double-stranded DNA. [0003] JC virus (JCV) is an opportunistic infectious pathogen, and the seropositive rate in the normal population is as high as 80%. JCV can be transmitted from mother to child through childbirth (placenta), breast-feeding or long-term co-living contact, and can also be transmitted through the respiratory and digestive tracts. After infection, it mostly exists in a latent state, and can be reactivated when the host's immune function is low...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68
CPCC12Q1/701C12Q1/6851C12Q1/686C12Q2545/101C12Q2561/101C12Q2521/531C12Q2563/143
Inventor 叶锋刘明坤余荣
Owner BEIJING SEARCH BIOTECH
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