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Nucleic acid detection method

A detection method and nucleic acid technology, applied in the field of molecular biology, can solve the problems of high environmental requirements such as detection temperature, long detection time, and high detection cost

Pending Publication Date: 2022-05-13
上海万子健生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, the traditional nucleic acid detection method has a long detection time, high requirements on the detection temperature and other environments, and requires the use of specific detection instruments, so the detection cost is high

Method used

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  • Nucleic acid detection method
  • Nucleic acid detection method
  • Nucleic acid detection method

Examples

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Embodiment 1

[0025] This embodiment provides a rapid nucleic acid detection method based on PCR and colloidal gold technology. The method combines the principles of PCR technology and colloidal gold technology to design a nucleic acid detection kit for human papillomavirus (HPV) type 18 , it can realize that after PCR amplification of HPV-18 virus nucleic acid DNA template, the product is detected and interpreted with a gold bar. The detection process includes steps:

[0026] 1) Amplify the pseudovirus synthetic template sequence of HPV-18 type virus L1 gene with a primer with a marker and another primer with a marker (see Table 1 for the specific primer sequence), the PCR system is as shown in Table 2, and the PCR The procedure is shown in Table 3, and the labeled HPV-18 double-stranded DNA product was obtained.

[0027] Table 1 PCR primers

[0028]

[0029] Table 2 PCR system preparation

[0030] component 25μL system 2x Taq Mix 12.5μL template 5μL F+...

Embodiment 2

[0036] This embodiment provides a rapid nucleic acid detection method based on RPA and colloidal gold technology, which combines the principles of RPA (Recombinase polymerase amplification) technology and colloidal gold technology, and designs a method for human papillomavirus (HPV) type 26 The nucleic acid detection kit of the HPV-26 virus can realize the isothermal amplification of the HPV-26 virus nucleic acid DNA template, and the product is detected and interpreted with a gold label. The detection process includes steps:

[0037] 1) Amplify the pseudovirus synthesis template sequence of the HPV-26 virus L1 gene and the pseudovirus synthesis of the internal reference β-actin gene with a labeled primer and another labeled primer (see Table 4 for specific primer sequences) The template sequence, isothermal amplification system is shown in Table 5, and the isothermal amplification procedure is shown in Table 6, to obtain a double-stranded DNA product with labeled HPV-26 and β...

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Abstract

The invention discloses a nucleic acid detection method which is characterized by comprising the following steps: 1) amplification of a to-be-detected nucleic acid sequence: a forward primer and a reverse primer are both specific amplification primers with markers, an amplification product is a double-stranded DNA product, and the two ends of the double-stranded DNA product are both provided with markers; (2) detecting the amplified double-stranded DNA product: a used detection tool is a chromatographic test strip, the chromatographic test strip contains a color developing substance, the color developing substance is attached with a pairing marker capable of being combined with the marker in the step (1), the detection temperature is room temperature, and a detection result is judged through color development of the chromatographic test strip; a color developing substance line, a target detection line and a quality control detection line are sequentially formed on the chromatographic test strip from a sample end to a handle end; only when the target detection line and the quality control detection line both develop colors, the result is judged to be positive. The method is short in detection time, low in cost, high in accuracy and convenient to operate, and a specific detection instrument is not needed.

Description

technical field [0001] The present invention relates to the technical field of molecular biology, in particular to a nucleic acid detection method. Background technique [0002] Polymerase chain reaction (Polymerase Chain Reaction, PCR) is a molecular biology technique used to replicate and amplify specific DNA fragments in vitro. In 1983, Mullis first proposed the idea, and in 1985 invented the polymerase chain reaction. PCR principle: DNA is denatured in vitro at 95°C to form a single strand; at ~60°C, primers and single strands are combined according to the principle of base complementary pairing; DNA polymerase moves along phosphate to five-carbon sugar (5'- 3') to synthesize complementary strands, so that the product obtained by PCR is usually double-stranded DNA. [0003] Isothermal Amplification Technology is also a kind of nucleic acid in vitro amplification technology. The reaction process is always maintained at a constant temperature, and the purpose of rapid nu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6804C12Q1/6844C12Q1/686
CPCC12Q1/6804C12Q1/6844C12Q1/686C12Q2563/131C12Q2565/625
Inventor 方剑秋沙海天袁梦瑶白艳军
Owner 上海万子健生物科技有限公司
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