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A novel kit and method for typing nine human papillomavirus subtypes

A typing method and kit technology, which is applied in the field of molecular biotechnology and genetic testing, can solve the problems of complex detection, time-consuming and labor-intensive, etc., and achieve the effect of simple result interpretation, low cost, and low requirements for facility conditions

Active Publication Date: 2018-08-03
广州市宝创生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, because it involves the distinction of multiple types, the detection is complicated, time-consuming and labor-intensive
[0006] At the same time, there is no human papillomavirus typing method in the market that combines the algorithm of mathematical permutation and combination with the detection method of nano-gold probes. Only ordinary PCR cycle instruments can be used to type more The number of human papillomavirus subtypes, such as 9 subtypes of human papillomavirus subtypes can be achieved with 5 tube detection reactions

Method used

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  • A novel kit and method for typing nine human papillomavirus subtypes
  • A novel kit and method for typing nine human papillomavirus subtypes
  • A novel kit and method for typing nine human papillomavirus subtypes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] The typing kit of embodiment 1 9 kinds of HPV hypotypes

[0040] A typing kit for 9 HPV subtypes, including the following four detection tubes and a control tube, wherein,

[0041] The first tube contains 4 types of detection primers and probes, specifically detection probes containing 11, 16, 31 and 42; the second tube contains 4 types of detection primers and probes, and the Only one type of detection probe is the same as the type in the first tube, specifically, the second tube contains detection probes of 6, 16, 33, and 39;

[0042] The third tube contains 3 types of detection primers and probes, only one type of the detection probe in the third tube is the same as the type in the first tube, and only one type is the same as the type in the second tube Same, specifically, the third tube contains 18, 31, 33 detection probes;

[0043] The fourth tube contains 3 types of detection primers and probes, only one type of the detection probe in the fourth tube is the same...

Embodiment 2 Embodiment 1

[0050] Example 2 The detection specificity of the 9 kinds of HPV subtypes described in Example 1 to distinguish 9 kinds of human papillomavirus subtypes

[0051] In this example, the primers and probes in Example 1 were used to prepare the reaction solution, with a total volume of 20 μL. Each sample was divided into 5 tubes for detection, and the detection components of each tube were: 1 μM primer, 1 μM probe and Hairpin Probe, 1μLAuNP-1 and AuNP-2, 0.5U Taq DNA polymerase, 2U endonuclease A, 0.2mM dNTP, were added to the system the corresponding target molecule of human papillomavirus subtype, the concentration is 10 4 Copy, the β-actin fragment template added in the last tube, the reaction system is composed of a reaction program: 94°C, 2min; 94°C, 5s, 72°C, 40s, 35 cycles; 72°C, 2min; 63°C, 10min; 55 ℃, 30min; 10℃, 2min.

[0052] The steps of detecting different subtypes of human papillomaviruses in the method of the present invention mainly include the following three sta...

Embodiment 3

[0054] Example 3 The detection sensitivity of the typing kit detection reaction of 9 kinds of HPV subtypes to distinguish 9 kinds of human papillomavirus subtypes

[0055] In this example, the primers and probes in Example 1 were used to prepare the reaction solution, the total volume was 20 μL, and the detection components in each tube were: 1 μM primer, 1 μM probe and Hairpin probe, 1 μL AuNP-1 and AuNP-2 , 0.5U Taq DNA polymerase, 2U endonuclease A, 0.2mM dNTP, add 10 5 、10 4 、10 3 、10 2 , 10, 1, and 0 copies of different subtypes of target molecules to be tested, tube 1 was added with different concentrations of template molecules of 11, 16, 31, and 42 subtypes; tube 2 was added with 6, 16, 33, and 39 different subtypes template molecules at different concentrations; tube 3 was added with different concentrations of template molecules of 18, 31, and 33 subtypes; tube 4 was added with different concentrations of template molecules with 35, 39, and 42 subtypes. The react...

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Abstract

The invention relates to a kit and a method for distinguishing more HPV (human papilloma virus) subtypes through a small number of detection reaction tubes by combining sequence combination with a nano-gold probe detection method. By a specific sequence combination, 9 typed human papilloma virus subtypes are distinguished by 4 detection tubes. By combining a mathematic permutation and combination algorithm with a molecular diagnostic technique, more human papilloma virus subtypes are distinguished by the smallest number of reaction tubes without increasing detection reaction flux, and detection operation difficulty and detection cost are reduced.

Description

technical field [0001] The invention belongs to the fields of molecular biotechnology and gene detection, in particular to a kit and a method for typing nine human papillomavirus subtypes. Background technique [0002] Human papillomavirus (Human Papilloma virus, HPV) belongs to the Papillomaviridae family, is a small molecule, non-encapsulated circular double-stranded DNA virus, the genome 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, and can only infect human skin and mucosal epithelial cells, causing various papillomas or warts on human skin and hyperplastic lesions of the genital tract epithelium. [0003] Human papillomaviruses th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/6858C12R1/93
CPCC12Q1/708C12Q2600/166
Inventor 王建平
Owner 广州市宝创生物技术有限公司
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