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Lentiviral gene transfer vector, preparation method and application thereof

A technology of gene transfer and lentivirus, which is applied in the field of genetic engineering, can solve problems affecting research and product development in the field, and there is no standardized and unified technology, so as to achieve the effect of stable protein expression, increased transfection efficiency, and increased expression

Inactive Publication Date: 2010-05-12
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, there is no commercialized EIAV gene transfer vector in China, and there is no standardized and unified technology, which affects research and product development in related fields

Method used

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  • Lentiviral gene transfer vector, preparation method and application thereof
  • Lentiviral gene transfer vector, preparation method and application thereof
  • Lentiviral gene transfer vector, preparation method and application thereof

Examples

Experimental program
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Embodiment 1

[0035] Example 1. Construction of pcPPTWCAG.

[0036] 1. PCR amplified EIAV donkey white blood cell attenuated vaccine strain (according to the "Technical Specifications for the Prevention and Treatment of Equine Infectious Anemia" published by the China Veterinary Drug Administration and the veterinary biological product "3413 Equine Infectious Anemia Agar Diffusion Test Antigen and Negative and Positive Serum ", "34140 Equine Infectious Anemia ELISA Antigen, Enzyme-labeled Antibody", "34141 Equine Infectious Anemia Complement Fixation Test Antigen" and "34142 Equine Infectious Anemia Virus Monoclonal Antibody-Enzyme Conjugate" and other reagent separation For the genome sequence of the virus, refer to "Yang Zhibiao, Wang Liu, Tong Guangzhi, Kong Xiangang, Qiu Huaji, Equine Infectious Anemia Virus Donkey Leukocyte Attenuated Vaccine Strain Provirus Whole Genome Nucleotide Sequence Analysis, Chinese Journal of Preventive Veterinary Medicine, 2001, 23( 3), the 3'LTR of 161-166"...

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PUM

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Abstract

The invention discloses an equine infectious anemia virus (EIAV) gene transfer vector, a constructed method and an application thereof. The vector of the invention comprises a CMV / R / U5 promoter, a 5' non-translated region homing sequence, a partial 5' gag gene coding region sequence, a central polypurine sequence, an Rev reaction element of EIAV, a CMVIE / chicken beta-actin promoter, a reporter gene, WPRE of WHBV, a polyclone locus, a poly (A) signal before EIAV 3'LTR, complete 3'LTR and partial sequence of eukaryon expression vector. The vector of the invention possesses common advantages of lentiviral vector, lacks auxiliary genes of all the wild-type virus, can carry one or a plurality of selected genes to transmit to target cells, can insert with target DNA sequence with large fragments and can lead the foreign genes and reporter genes to express at a high level in the cells.

Description

technical field [0001] The present invention relates to a lentiviral gene transfer vector, in particular to a defective lentiviral gene transfer vector derived from Equine Infectious Anemia Virus (EIAV) and its construction method. The application in gene transfer of cells in vitro belongs to the field of genetic engineering. Background technique [0002] The currently constructed EIAV gene transfer vectors are mainly derived from American strains reported abroad, and there are large differences between American strains and Chinese strains, with nucleotide homology between 78.4% and 80%. There is no related report of the same type in China, and no one has used woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) and chicken β-actin promoter (CAGp) to enhance the expression of the reporter gene on the EIAV gene transfer vector. At present, there is no commercialized EIAV gene transfer vector in China, and there is no standardized and unified technology, w...

Claims

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Application Information

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IPC IPC(8): C12N15/86C12N5/10C12N7/01
Inventor 韩凌霞童光志曲连东司昌德于海波孟庆文姜骞刘家森孙成群
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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