Preparation method of HIV-1 nucleic acid detection quality control product based on influenza virus vector

Abstract

The invention provides a preparation method of an HIV-1 nucleic acid detection quality control product based on an influenza virus vector. The quality control product is obtained by performing multiplication culture, inactivation and purification on a recombinant influenza virus carrying an HIV-1 conservative gene. The preparation method of the HIV-1 nucleic acid detection quality control productbased on the influenza virus vector comprises the following steps: (1) constructing a CRISPR-Cas9 large fragment knock-in system which is inserted into an HIV-1 conservative gene and can be used for saving influenza viruses PR8; (2) co-transfecting 293T cells with the constructed system, PR8 influenza viruses PB1, PB2, PA, HA, NP, NA and M protein encoding plasmids, and exciting functions to saveinfluenza viruses, so as to obtain recombinant influenza viruses with HIV-15-'UTR; and (3) performing multiplication culture, inactivation and purification on the constructed influenza virus to obtainthe HIV-1 nucleic acid detection quality control product. The kit has the characteristic of highly simulating the HIV-1 virus, and can perform comprehensive detection on HIV-1 nucleic acid detection;The whole preparation process is simple and convenient, and the HIV-1 nucleic acid detection quality control product is low in cost and easy to store and transport, and has a good application prospect.

Description

technical field [0001] The invention belongs to the field of HIV-1 nucleic acid detection and specifically designs a method for preparing a HIV-1 nucleic acid detection quality control product based on influenza virus vectors. Background technique [0002] Human immunodeficiency virus (Humanimmunodeficiencyvirus, HIV) is an RNA virus that is transmitted through blood, body fluids, etc. It is a retrovirus and is the chief culprit of AIDS. HIV-1 virus is the main subtype prevalent in my country at present, accounting for more than 90%. Although China's AIDS epidemic is still at a relatively low level overall, the number of AIDS patients is increasing year by year. In recent years, my country has reported more than 100,000 newly discovered HIV-infected persons / AIDS patients every year, and it has shown an increasing trend every year. As of 2018, the number of surviving AIDS patients and infected persons nationwide has exceeded 820,000. Therefore, Accurate, efficient and accura...

AI Technical Summary

Problems solved by technology

[0004] At present, HIV-1 nucleic acid detection quality control products mainly include serum / plasma from positive patients, artificially prepared pseudovirion particles, or directly using naked RNA fragments, all of which have their own limitations: quality control from positive patients The nucleic acid load of the product is unstable, the difference between preparation batches is not easy to control, and has biological safety hazards; the naked RNA itself is very poor in stability and is easily affected by the environment, which has an incalculable impact on the final test results; Virus particles generally use in vitro recombination technology, and use phospholipid bilayer structure to report RNA fragments, such as phage particles, which are highly imitative, but there are still defects such as high preparation costs and technical thresholds, and it is difficult to use large-scale in actual work. Controls using this source

[0005] Influenza virus is currently considered to be an extremely ideal viral gene manipulation carrier. In the past, due to the complexity of editing and preparing specific genotype influenza virus technology, it was difficult to apply to industrial production.

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