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Nucleotide sequences of primers for skin HPV type detection based on constant-temperature nucleic acid amplification technology and application thereof

A nucleotide sequence and nucleic acid sequence technology is applied in the field of primers for skin type papillomavirus typing and detection, which can solve the problems of long time period, large total amount of HPV types, unfavorable technology promotion and application, etc. Sensitivity and specificity of reaction, less false positive reaction, rapid response effect

Active Publication Date: 2019-09-03
THE FIRST HOSPITAL OF CHINA MEDICIAL UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method also has obvious disadvantages, which are mainly manifested in: 1) The PCR reaction only uses two specific primers (upstream and downstream DNA primers) for amplification, and the total amount of HPV types is huge. have limited specificity and are prone to false positive reactions
2) The PCR reaction requires a precise temperature control system to meet the control of its variable temperature reaction conditions, and the reading and interpretation of the PCR amplification results requires professional fluorescence detection equipment and personnel reserves with professional skills, which greatly increases the The cost of equipment and personnel for PCR typing and identification is not conducive to the promotion and application of technology
3) The PCR typing reaction requires a long period of time (sample processing, DNA extraction, and PCR reaction can take more than 5-6 hours), which is not conducive to the rapid clinical detection of HPV typing needs

Method used

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  • Nucleotide sequences of primers for skin HPV type detection based on constant-temperature nucleic acid amplification technology and application thereof
  • Nucleotide sequences of primers for skin HPV type detection based on constant-temperature nucleic acid amplification technology and application thereof
  • Nucleotide sequences of primers for skin HPV type detection based on constant-temperature nucleic acid amplification technology and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Primer nucleic acid sequence design and screening.

[0027] 1. Primer nucleic acid sequence design method: the basic structure of CAMP primer is as follows: figure 1 As shown, the complementary pairing patterns of the three sets of basic primers of CAMP primers and template DNA are as follows figure 1 shown. The NF primer is formed by linking N with F1 (5'-N-F1-3'), and the NR primer is formed by linking Nc with R1 (5'-Nc-R1-3'). Based on this principle, the primers for CAMP typing and identification of 8 common HPVs were designed. The HPV-L1 gene sequences corresponding to all CAMP primers were queried and downloaded from the NCBI-Nucleotide database.

[0028] 2. The specific screening method for the nucleic acid sequence of the primer comprises the following steps.

[0029] 1) Screening method for primer nucleic acid sequences: Summarize the L1 gene sequences of all common skin-type HPVs and conduct multiple sequence comparison analysis. The CAMP primer...

Embodiment 2

[0032] Example 2 HPV type detection was performed on clinical samples.

[0033] The skin scrapings from the skin lesions of patients with clinical plantar warts or common warts were scraped, and the total DNA of the skin scraping samples was extracted, which was used as the DNA sample to be tested for primer amplification identification. The experimental group and the control group were set up the same as before, and the CAMP primers screened out by the 8 skin-type HPVs involved in the present invention were used to detect the same sample, and the amplification results of the CAMP primers of different clinical samples were recorded. At the same time, corresponding DNA samples of skin lesions were amplified and sequenced using HPV universal primers to clarify the specific HPV infection type of the samples. Compare the CAMP amplification results and HPV sequencing results of the same sample to judge the accuracy of the screened primers in actual clinical testing.

[0034] Using...

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Abstract

The invention relates to the fields of DNA amplification and biological sample detection, and in particular, relates to eight primers for skin papillomavirus type detection based on a constant-temperature nucleic acid amplification technology. The eight pairs of primers can be used for preparing diagnostic products, such as kits, for rapid detection of skin-type HPV types. The HPV type identification primers involved in the invention are all precisely designed and screened by a large number of experiments and then determined, and have quite high reaction sensitivity and specificity. Finally, the primers pass inspection by clinical sample amplification test and are the best selection of the 8-type HPV type detection primers based on the CAMP technology. Nucleotide sequences can quickly andaccurately detect the type of skin-type HPV, and can meet the clinical needs of the skin-type HPV types.

Description

technical field [0001] The invention relates to the field of DNA amplification and biological sample detection, in particular to eight kinds of primers for skin type papillomavirus type detection based on constant temperature nucleic acid amplification technology. Background technique [0002] Human papillomavirus (HPV) is a small virus with double-stranded circular DNA as its genetic material, and its genome consists of about 8000 base pairs. HPV virus particles are about 50nm in diameter and belong to the family Polyomavacuoviridae. Clinically, HPV can be roughly divided into two main types according to the types of HPV-infected tissues, namely, skin type and mucosal type. HPV types, and new HPV types are constantly being discovered. Clinically, HPV infection can cause various types of diseases, among which benign diseases include skin warts, verrucous epidermal dysplasia, cervical CIN, etc.; malignant diseases include solar keratosis, non-melanoma skin cancer, head and ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6844C12N15/11
CPCC12Q1/708C12Q1/6844C12Q2531/119
Inventor 孙宇哲高兴华毛瑞齐瑞群杜昱光梁祖源陈洪铎
Owner THE FIRST HOSPITAL OF CHINA MEDICIAL UNIV