Typing detection method and kit for high-risk human papilloma virus (HPV)

A technology of human papillomavirus and detection method, which is applied in the field of high-risk human papillomavirus typing detection methods and kits, and can solve the problems of restriction of types of gene sequence detection and the like

Inactive Publication Date: 2017-08-15
SHANGHAI XINGYAO MED TECH DEV CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Single-probe real-time quantitative PCR, because each probe is designed for a specific target site, with high specificity, each probe can only detect one target sequence, if multiple target sequences need to be detected, Multiple probes need to be added. Due to the limitation of the absorption wavelength of the fluorescent label of the probe, the types of gene sequence detection are restricted.

Method used

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  • Typing detection method and kit for high-risk human papilloma virus (HPV)
  • Typing detection method and kit for high-risk human papilloma virus (HPV)
  • Typing detection method and kit for high-risk human papilloma virus (HPV)

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Effect test

Embodiment 1

[0123] Embodiment 1 Human papillomavirus 16, 18, 31, 52, 59 type nucleic acid typing detection kit (fluorescent PCR method)

[0124] 1. Primer Probe Synthesis

[0125] Entrust Yingwei Jieji (Shanghai) Trading Co., Ltd. to synthesize the primer probe. The sequence of the synthesized primer probe is as follows:

[0126] The upstream primers are:

[0127] 5'-CAACTATTTGTTACTGTTGTTGATACTAC-3' as shown in SEQ ID No.1;

[0128] 5'- CAATTATTTGTTACTGTGGTAGATACCAC -3' as shown in SEQ ID No.2;

[0129] 5'-CAGTTGTTTGTCACAGTTGTGGATACCAC-3' as shown in SEQ ID No.3;

[0130] 5'-CAATTGTTTTTAACAGTTGTAGATCCTAC-3', as shown in SEQ ID No.4;

[0131] Downstream primers are:

[0132] 5'-GAAATATAAACTGCAAATCAAATTCCTC-3', as shown in SEQ ID No.5;

[0133] 5'- GAAAAATAAATTGTAAATCAAATTCCTC-3', as shown in SEQ ID No.6;

[0134] 5'-GAAATATAAATTGTAAATCAAATTCCTC-3', as shown in SEQ ID No.7;

[0135] 5'- GAAAAATAAACTGCAAATCATATTCCTC-3', as shown in SEQ ID No.8;

[0136] 5'-GAAAAATAAACTGTAAATCATATTCC...

Embodiment 2

[0165] Application of embodiment 2 human papillomavirus 16, 18, 31, 52, 59 type nucleic acid typing detection kit (fluorescent PCR method)

[0166] Using the kit obtained in the present invention to test 20 cases of clinical samples, and at the same time use the human papillomavirus genotyping (type 25) detection kit (PCR-reverse dot hybridization method) produced by Aikang Biotechnology Co., Ltd. for parallel comparison Test and evaluate the effect of the kit.

[0167] 1. Human papillomavirus 16, 18, 31, 52, 59 nucleic acid typing detection kit (fluorescent PCR method) for detection of clinical samples

[0168] (1) Reagent preparation

[0169] Prepare the reaction solution according to the number of samples n (number of samples = number of samples to be tested + 3 reference substances + 1):

[0170] Take PCR buffer n×9 μL, enzyme mixture n×3 μL, primer probe n×8 μL in a centrifuge tube and mix evenly; centrifuge at low speed for a few seconds, and dispense 20 μL / tube into r...

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Abstract

The invention provides a typing detection method and kit for a high-risk human papilloma virus (HPV). According to the method, a method for performing fluorescence nucleic acid amplification by adopting multiple pairs of primers and multiple probes is combined with fusion curve analysis, so that multiple target nucleic acid sequences can be detected in a single-wavelength fluorescence channel; an L1 segment sequence of an HPV genome is used as a target sequence for designing an HPV specific primer, a TaqMan probe and a PCO hybridization probe which are not completely complementary with the TaqMan probe, and a 3' end of the hybridization probe can be subjected to fluorescence quenching group labeling or non-fluorescence quenching group labeling according to purposes and cannot be extended after the hybridization probe is labeled, so that an HPV gene typing kit containing a primer probe and other fluorescence PCR components is researched; and by combining fluorescence PCR amplification with single-wavelength fusion analysis, the types 16, 18, 31, 52 and 59 of the HPV can be identified.

Description

technical field [0001] The invention relates to a high-risk human papilloma virus typing detection method and kit, which belong to the field of biotechnology. Background technique [0002] Human papillomavirus (Human papillomavirus, HPV) belongs to the Papillomaviridae family, is a small molecule, non-encapsulated, circular double-stranded DNA virus, the genome length is about 8000 base pairs (bp), divided into 3 There are three functional regions, namely early transcribed region (E region), late transcribed region (L region) and non-transcribed region (long control region, LCR). HPV infects humans through direct or indirect contact with contaminated objects or through sexual transmission. The virus is not only host-specific, but also tissue-specific. It can only infect human skin and mucosal epithelial cells, and can cause squamous epithelial proliferation of human skin and mucous membranes. More than 130 kinds of papillae that cause human skin have been isolated so far. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
CPCC12Q1/686C12Q1/705C12Q2561/101
Inventor 郭圣明吴大治夏懿
Owner SHANGHAI XINGYAO MED TECH DEV CO LTD
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