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Method for preparing recombinant avian adeno-associated virus and use of recombinant avian adeno-associated virus

A recombinant baculovirus and virus technology, applied in the field of viruses, can solve the problems of reducing the scope of use of recombinant avian adenoviruses, low titer of recombinant avian adenoviruses and the like

Inactive Publication Date: 2017-04-26
JIANGSU AGRI ANIMAL HUSBANDRY VOCATIONAL COLLEGE
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Problems solved by technology

[0004] There are many methods for preparing recombinant adeno-associated virus, but the virus titer of recombinant adeno-associated virus prepared by these methods is low, which greatly reduces the scope of use of recombinant adeno-associated virus

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  • Method for preparing recombinant avian adeno-associated virus and use of recombinant avian adeno-associated virus
  • Method for preparing recombinant avian adeno-associated virus and use of recombinant avian adeno-associated virus
  • Method for preparing recombinant avian adeno-associated virus and use of recombinant avian adeno-associated virus

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Embodiment Construction

[0041] The present invention will be described in further detail below in conjunction with the accompanying drawings and embodiments, so that those skilled in the art can implement it with reference to the description.

[0042] It should be understood that terms such as "having", "comprising" and "including" used herein do not exclude the presence or addition of one or more other elements or combinations thereof.

[0043] 1. Using recombinant DNA technology, prepare three recombinant baculovirus transfer vectors, which are respectively marked as pFastBac-AITR-LYZ, pFastBac1-VP, and pFastBac-Dual-Rep;

[0044] Using DNA recombination technology, on the basis of pAITR-KLKI and p215C3LYZ, the human lysozyme gene expression cassette controlled by the chicken ovalbumin gene promoter was inserted between the left and right ITRs of AAAV, and the recombinant vector pFastBac- AITR-LYZ, the construction idea is as follows figure 1 As shown, the identification diagram is shown in figu...

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Abstract

The invention discloses a method for preparing a recombinant avian adeno-associated virus. The method comprises 1, preparing three recombinant baculovirus transfer vectors by a DNA recombination technology, 2, respectively carrying out transposition on the three recombinant baculovirus transfer vectors to a DH10Bac competent cell, carrying out screening, and extracting three recombinant shuttle vector plasmid DNAs through an alkaline lysis method, 3, uniformly mixing the three recombinant shuttle vector plasmid DNAs and a transfection reagent PEI according to a certain proportion, transfecting a Sf9 insect cell with the mixture, and after transfection for three days, preparing the recombinant avian adeno-associated virus expressing a human lysozyme. The invention also provides the use of the recombinant avian adeno-associated virus expressing a human lysozyme. The titer of the recombinant avian adeno-associated virus is ten times that of a recombinant avian adeno-associated virus obtained by the conventional method. The human lysozyme expressed by the recombinant avian adeno-associated virus can produce antibacterial effects on the common pathogens.

Description

technical field [0001] The invention relates to the technical field of viruses, more specifically, the invention relates to a preparation method and application of a recombinant adeno-associated virus. Background technique [0002] Lysozyme is a natural antibacterial agent. Lysozyme not only has a direct bacteriolytic effect on many Gram-positive bacteria, but also has a certain inhibitory effect on Gram-negative bacteria and some fungi. It also has a certain inhibitory effect. At the same time, human lysozyme is non-toxic to humans and animals, will not cause public health problems caused by drug residues, has no adverse reactions and side effects on the human body, and will not produce drug resistance even if it is used for a long time. [0003] The baculovirus expression system is a eukaryotic expression system developed in the 1980s. It is one of the four major expression systems in genetic engineering. It has the advantages of high expression of foreign genes, high bio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N9/36C12N15/866A61K38/47A61P31/04A61P31/10
CPCC12N7/00A61K38/47C12N9/2462C12N15/86C12N2710/14043C12N2750/14121C12N2750/14151C12Y302/01017
Inventor 王永娟左伟勇洪伟鸣朱善元
Owner JIANGSU AGRI ANIMAL HUSBANDRY VOCATIONAL COLLEGE
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