Construction technology of recombinant insect cross virus expression vector

A technology of expression vectors and viruses, which is applied in the field of construction of new insect recombinant hybrid virus expression vectors, can solve the problems of long time for viralization, small host insects, and difficult industrialization level, and achieve a wide range of hosts and industrialization The effect of level development and production level improvement

Inactive Publication Date: 2002-02-20
ZHEJIANG UNIV
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Problems solved by technology

However, the BmNPV expression system also has disadvantages: because BmNPV host is specific and only has the ability to infect silkworms, only silkworm cultured cells can be used in the construction of recombinant viruses, and silkworm cells have the characteristics of slow growth and subculture, so It takes a long time for the construction of recombinant virus, screening and the multiplication of a large number of viruses for expression
But its shortcomings are also obvious, that is, the host insects are small and difficult to raise, and it is difficult to be suitable for the development of industrialization level.

Method used

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  • Construction technology of recombinant insect cross virus expression vector
  • Construction technology of recombinant insect cross virus expression vector
  • Construction technology of recombinant insect cross virus expression vector

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Embodiment Construction

[0021] 1. Materials: Insect cultured cells Sf21, Tn-5 and silkworm BmN cells were subcultured and preserved by the Bioengineering Laboratory of the Silkworm Department of the School of Animal Science, Zhejiang University. AcNPV and BmNPV viruses were preserved by the Silkworm Physiological and Pathological Laboratory of the same university. Sf21 cell culture medium TC-100 is a product of GibcoBRL, add 10% fetal bovine serum (v / v) before use, Tn-5 cells use serum-free ESF921 medium as a product of Expression systems, and the culture temperature is 27°C . Bombyx mori larvae are educated with artificial feed for 1-3 instars at 29°C, and with mulberry leaves for 4-5 instars at 23-25°C. Various enzymes and PCR amplification kits used in gene manipulation are products of Takara Biomedicals, Nippon Takara Shuzo. The TA cloning kit, plasmid pCR2.1 and insect baculovirus transfer plasmids pBlueBacHisA, B, C were purchased from Invitrogen Company of the United States, and the insect ba...

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Abstract

The present invention relates to construction technique of insect recombination hybridization virus expression carrier, it uses isogenous recombination priciple to construct hybridization reconstruction virus expression carrier HyNPV situated between AcNPV and BmNPV. Said expression carrier solves the problem of narrow host range of BmNPV expression system and degradation of recombination object product by protease. It possesses the advantage of broad host range, not only can infect domestic silkworm, but also can infect commonly used Sf21, Sf9 and Tn-5 culture cell, also can use Sf21, Sf9 culture cell to instead domestic silkworm cell to hasthen the preparation of recombination virus.

Description

technical field [0001] The invention relates to a construction technology of a novel insect recombinant hybrid virus expression vector. Background technique [0002] The production of recombinant proteins by means of genetic engineering such as recombinant DNA is the main driving force behind the rapid development of the biopharmaceutical industry in recent years. At present, there are mainly three different levels of expression systems for the production of recombinant proteins: prokaryotic, insect and mammalian cells. Among them, the insect baculovirus expression system Baculovirus Expression Vector System, referred to as BEVS, has become the most effective due to its high-efficiency expression, relatively complete protein post-translational processing, large-scale expression production using individual insects or their cultured cells, and good safety. One of the eukaryotic expression systems. At present, there are mainly two types of BEVS, Autographa californica nuclear...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09C12N15/63C12N15/64C12N15/86
Inventor 吴小锋
Owner ZHEJIANG UNIV
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