Novel kit and method for typing 21 human papilloma virus subtypes

A typing method and kit technology, applied in the field of molecular biotechnology and genetic testing, can solve the problems of complex testing, time-consuming and labor-intensive, and achieve the effects of stable storage conditions, low cost, and reduced complexity and testing costs.

Active Publication Date: 2017-09-05
广州市宝创生物技术有限公司
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  • Abstract
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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 present, 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 7 tube detection reactions to achieve 21 subtypes of human papillomavirus typing

Method used

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  • Novel kit and method for typing 21 human papilloma virus subtypes
  • Novel kit and method for typing 21 human papilloma virus subtypes
  • Novel kit and method for typing 21 human papilloma virus subtypes

Examples

Experimental program
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Embodiment 12

[0036] The typing kit of embodiment 1 21 kinds of HPV hypotypes

[0037]A typing kit for 21 HPV subtypes, including the following 6 detection tubes and 1 control tube, wherein,

[0038] The first tube contains 6 types of detection primers and probes, specifically 11, 16, 31, 35, 59, and 68 detection primers and probes;

[0039] The second tube contains 6 types of detection primers and probes, and only one type of detection primers and probes in the second tube is the same as the model in the first tube. Specifically, the second tube contains 6, 16 , 52, 53, 56, 68 detection primers and probes;

[0040] The third tube contains 6 types of detection primers and probes. Only one type of the detection primers and probes in the third tube is the same as that in the first tube, and only one type is the same as that in the second tube. The models are the same, specifically, the third tube contains detection primers and probes of 31, 33, 45, 52, 58, and 81;

[0041] The fourth tube ...

Embodiment 2 Embodiment 1

[0072] Example 2 The seven-tube detection reaction of the 21 HPV subtype typing kits described in Example 1 distinguishes the detection specificity of 21 HPV subtypes

[0073] In this example, the primers and probes in Example 1 were used to prepare the reaction solution, the total volume was 20 μL, and each sample was divided into 7 tubes for detection, and the detection components of 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, were added to the system the target molecules corresponding to the subtype of human papillomavirus, the concentration is 10 4 Copy, add β-actin fragments to the last tube, and 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°C, 30min; 10°C, 2min.

[0074] The detection method of the present invention utilizes a new type of in vitro diagnostic method (nano-gold visual detection met...

Embodiment 3 7

[0076] Embodiment 3 Seven-tube detection reaction distinguishes the detection sensitivity of 21 kinds of human papillomavirus subtypes

[0077] In this example, the primers and probes in Example 1 were used to prepare the reaction solution, the total volume was 20 μL, and each sample was divided into 7 tubes for detection, and the detection components of 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 18, 33, 56, 83, 59, and 68 subtypes; tube 2 was added with 31, 39, 42, Template molecules of different concentrations of 43, 82, and 68 subtypes; tube 3 was added with different concentrations of template molecules of 16, 35, 43, 53, 59, and 66 subtypes; tube 4 was added with 6, 35, 42, 45, 56, Template molecules...

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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, 21 typed human papilloma virus subtypes are distinguished by 6 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 field of molecular biotechnology and gene detection, in particular to a kit and method for typing 21 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 that in...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
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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