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Recombined rhabdovirus AcBac delt CC-GP41 and constructing method thereof

A technology of recombinant baculovirus and CC-GP41, which is applied in the field of bioengineering technology and medicine, to achieve the effects of large expression, convenient expression and subsequent purification, and simple construction and operation

Inactive Publication Date: 2008-01-30
WUHAN INST OF VIROLOGY CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, this invention is not a full-length GP41 protein, which is quite different from the natural GP41 protein

Method used

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  • Recombined rhabdovirus AcBac delt CC-GP41 and constructing method thereof
  • Recombined rhabdovirus AcBac delt CC-GP41 and constructing method thereof
  • Recombined rhabdovirus AcBac delt CC-GP41 and constructing method thereof

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Embodiment Construction

[0016] recombinant baculovirus AcBac △CC - GP41, deposit number: CCTCCNO.V200702.

[0017] Below in conjunction with accompanying drawing to recombinant baculovirus AcBac of the present invention △CC - The construction method of GP41 is further explained.

[0018] As can be seen from Figure 3, the recombinant baculovirus AcBac △CC - The construction method of GP41 is carried out in the following order of steps:

[0019] a, amplify the gp41 gene from HIV-1 virus with polymerase chain reaction (PCR);

[0020] b. According to the instructions of the pGEM-T Easy vector kit from Promega, USA, the amplified gp41 gene sequence was cloned into pGEM-T Easy, and passed through isopropyl-B-D thiogalactoside and 5-bromo-4-chloro -3-indole-β-D-galactoside (IPTG / X-gal) blue-white screening and restriction endonuclease EcoRI digestion identification, the plasmid pGEM-T-gp41 containing the gp41 gene was obtained and verified by sequencing No mutation;

[0021] c. Using the donor plasmid...

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Abstract

The invention provides a recombinant baculovirus AcBacDeltaCC-GP41 and an establishment method thereof and relates to the technical field of biological engineering technique and pharmacological. The recombinant baculovirus is CCTCC NO.V200702 and the establishment method comprises the steps: the expression system of Bac-to-Bac baculovirus developed by INVITROGEN Company is used, an AcMNPV baculovirus carrier is selected, then double-deletion shuttle carrier Ac-bacDeltaCC is obtained through a reformation of an AcMNPV shuttle carrier and last the recombinant baculovirus AcBacDeltaCC-GP41 is obtained by using insect cell expression system. The invention has the advantages that the virus is the protein recombinant virus which can high effectively express HIV-1 and has the full length of GP41 and the structure and function thereof are close to natural protein and beneficial for studying natural conformation of GP41 and paves the solid foundation for fully explaining mechanism of HIV-1 entering cells and the immune diagnose and vaccine development based on GP41 and the prevention and curing of AIDS.

Description

technical field [0001] The invention relates to the fields of bioengineering technology and medical technology, in particular to a recombinant baculovirus capable of highly expressing HIV-1 envelope glycoprotein GP41. The invention also relates to the construction method of the recombinant baculovirus. Background technique [0002] At present, the most widely used baculovirus expression vector is the commercialized baculovirus Bac-to-Bac system, and the most commonly used baculovirus vector is the A. californica polynuclear capsid nucleopolyhedrosis virus (AcMNPV). The Bac-to-Bac baculovirus expression system is mainly composed of donor plasmids, shuttle vector bacmid, and insect cells. Because this method is fast and convenient, the screening cycle of the entire recombinant virus is 7 to 10 days, and can be continuously screened. At the same time, Donor plasmids can construct more restriction enzyme sites and be modified to facilitate the insertion of foreign genes. [00...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/866
Inventor 苏正元李娟王华林邓菲胡志红袁丽
Owner WUHAN INST OF VIROLOGY CHINESE ACADEMY OF SCI
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