Method for flexibly detecting human papilloma virus genotypes in broad spectrum

Abstract

The invention provides a method for flexibly detecting human papilloma virus genotypes in a broad spectrum. The detected types contain 45 carcinogenic and pathogenic HPV genotypes (subtypes) described by the International Agency for Research on Cancer IARC and one giant wart-inducing type newly authenticated and reported by China CDC, and the coverage exceeds the sum of the types involved by the present various similar kits. According to the method provided by the invention, mass-spectrum scanning multiple PCR is taken as a platform, 21 carcinogenic and 2 high-incidence wart-inducing types with the highest clinic need frequency are detected as an independent reaction, and only two reactions are needed in total, thus further meeting the needs of clinic emphasis detection and difficult and complicated disease broad-spectrum screening; detection combinations are flexible and cost performance is high; several and even thousands of samples can be detected in one day; a result is automatically reported in real time, without the influences of human factors; tens of HPV types can be accurately detected simultaneously in one step. The method embodies the unique advantages of high flux, high flexibility, high automation and wide linear range, and provides an indispensable detection tool for comprehensive universal screening for pathogenic HPV infection.

Description

Technical field [0001] The invention relates to the field of detection and identification of pathogenic microorganism preparations, in particular to a broad-spectrum and flexible method for detecting the genotype of human papillomavirus. Background technique [0002] In 1983, German virologist Harald zur Hausen first discovered that some types of human papillomavirus (HPV) are related to cervical cancer. The research results make cervical cancer the only human cancer that can be prevented, detected early, and cured. In 2003, the World Health Organization’s International Oncology Research Agency (WHO_IARC) published an analysis report drawn by 11 research institutions in 9 countries. According to the relationship between different types of HPV and cervical cancer, 30 HPV genotypes will be detected. There are three major categories, including 15 high-risk carcinogenic types: 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73, 82 (W13B / MM4 subtype and IS39 subtype Type); 3 hig...

AI Technical Summary

Problems solved by technology

It cannot meet the current clinical needs for broad-spectrum screening of HPV, and its detection results will cause more than 1% of people to miss the diagnosis

[0009] 2. The detection sensitivity of the hybridization method is low, about 5000copies / ml, and visual inspection is required, and the detection linear range is narrow

Through comparative experiments, it was found that at present, more than 20 genotypes can be detected at one time. In actual testing, only about 3 high-load genotypes can be detected at most at one time, while high-risk and low-load genotypes are missed, such as After the HPV16 virus is integrated into the human genome, it is prone to strong carcinogenicity, but it is prone to missed detection due to the loss of the L1 gene

[0010] 3. The number of types detected by the real-time fluorescent PCR method is limited, and the typing kit with the highest number of types only has 13 types, and 5 reactions are required to complete the detection

[0011] 4. The current PCR-mass spectrometry method uses multiple PCR probes to mix and identify similar sequences in the amplification region of the same universal primer, which is prone to false positive problems of cross-mismatching, making it difficult to repeat the results

Using the double-gene probe method to detect 30 genotypes requires three reactions to be detected at the same time, and the type coverage is not comprehensive enough

In addition, due to the high cost, beyond the affordability of routine clinical testing charges, it is mainly suitable for paraffin samples that are difficult to accurately detect by other methods

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