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A kind of pseudovirus, its packaging method and drug evaluation system

A pseudovirus and packaging technology, applied in the field of genetic engineering, can solve the problems of unable to drive the expression of wild-type S protein, unable to make up for the lack of promoters, etc., to avoid secondary infection, improve infection ability, and increase the effect of expression.

Active Publication Date: 2022-05-03
中吉当康(北京)基因技术有限公司
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Problems solved by technology

However, in 2004, when scientists packaged the SARS-CoV pseudovirus system, they found that in the packaging cells, the CMV promoter could not drive the expression of wild-type S protein. Through codon optimization, the amount of S protein increased, but Still can't make up for the lack of promoters

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  • A kind of pseudovirus, its packaging method and drug evaluation system
  • A kind of pseudovirus, its packaging method and drug evaluation system
  • A kind of pseudovirus, its packaging method and drug evaluation system

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Embodiment Construction

[0047] In order to better explain the present invention and facilitate understanding, the present invention will be described in detail below through specific embodiments in conjunction with the accompanying drawings.

[0048] The main innovation of the present invention is: when packaging pseudoviruses, CBA is selected as the promoter to drive the expression of the envelope protein Spike, and the Spike DNA sequence optimized for human expression is used to replace the wild-type Spike DNA sequence, thereby obtaining high-expression Lentiviral pseudoviruses of the S envelope protein. When constructing the new coronary pneumonia drug evaluation system of the present invention, a monoclonal 293T cell line that can overexpress and stably express the hACE2 receptor was also constructed to amplify the infection effect of the pseudovirus on cells, which is helpful for the evaluation of drug efficacy using the system , more accurate evaluation results can be obtained. In the process ...

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Abstract

The present invention relates to a pseudovirus, which comprises: a plasmid expressing the envelope protein Spike, a reporter gene plasmid and a packaging auxiliary plasmid; the plasmid expressing the envelope protein Spike contains a CBA promoter and a human optimized Spike expression sequence, Used to drive the expression of the envelope protein Spike. The pseudovirus of the present invention can simulate the process of infecting cells with wild-type novel coronavirus, and can be used to study the relationship between virus and host cells, clone virus receptors, replace traditional neutralization tests, and be used for differential diagnosis of diseases and antiviral drugs The evaluation and screening of vaccines and the evaluation of vaccine immune effects. The present invention also relates to the packaging method of the pseudovirus and the new coronavirus drug evaluation system comprising the pseudovirus, wherein the pseudovirus is used to infect the 293T monoclonal cells overexpressing the human ACE2 gene under the condition of drug interference, and the pseudovirus is detected by detecting the pseudovirus. The expression level of the reporter gene in the virus (such as detection of fluorescence intensity, etc.), and then initially evaluate the effectiveness of the drug.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a pseudovirus, a packaging method thereof and a drug evaluation system. Background technique [0002] In the study of virus function, the most direct and efficient method is of course to manipulate wild-type live viruses. However, for highly pathogenic and infectious viruses such as the new coronavirus, the operation of wild-type live viruses needs to be carried out in biosafety protection (P3) laboratories above level 3. At present, there are only more than 40 P3 laboratories in China, and most of them Belongs to the CDC system, non-conventional scientific research purposes. In addition, in order to prevent biological hazards, the P3 laboratory has very strict protection conditions, which greatly reduces the operating efficiency. [0003] In order to be able to conduct research on the new coronavirus in the second-level biosafety laboratory, pseudovirus technology is a very ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/01C12N15/50C12N15/867C12N5/10C12Q1/02
CPCC12N7/00C07K14/005C12N15/86G01N33/5008C12N2770/20022C12N2740/15043C12N2770/20021
Inventor 董文吉赵忠亮刘子瑾张艳君程谟斌李昌锋
Owner 中吉当康(北京)基因技术有限公司
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