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Universal transfer vector for pseudorabies virus capable of independently expressing dual genes and construction method and application of universal transfer vector

A technology of pseudorabies virus and transfer vector, which is applied in the direction of using vector to introduce foreign genetic material, cells modified by introducing foreign genetic material, recombinant DNA technology, etc.

Inactive Publication Date: 2014-12-10
GUANGDONG WENS DAHUANONG BIOTECH +1
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is currently no universal transfer vector for pseudorabies virus

Method used

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  • Universal transfer vector for pseudorabies virus capable of independently expressing dual genes and construction method and application of universal transfer vector
  • Universal transfer vector for pseudorabies virus capable of independently expressing dual genes and construction method and application of universal transfer vector
  • Universal transfer vector for pseudorabies virus capable of independently expressing dual genes and construction method and application of universal transfer vector

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Embodiment 1

[0034] A pseudorabies virus universal transfer vector that independently expresses double genes, which is inserted into the XbaI site between gEL and gER in the pUC19 / HEK-gE plasmid vector containing gEL and gER, containing two CMV promoters, located at 2 A multiple cloning site downstream of a CMV promoter and a double gene expression cassette of a BGH pA signal sequence, the specific construction process is as follows:

[0035] 1) Obtain homologous recombination arms gEL and gER from PRV genomic DNA amplification by PCR method:

[0036] Referring to the published porcine pseudorabies virus genome sequence in GenBank, design the following three pairs of primers:

[0037] P1 (SEQ ID NO.2): 5-ATTGGTACCGCTGCTGTTCGTCTCGCG-3 is the upstream primer of gEL, and a Kpn Ⅰ site is introduced at the 5′-end;

[0038] P2 (SEQ ID NO.3): 5-GTCTCTAGAAGGGACTCGGGACCTCG-3 is the downstream primer of gEL, and Xba I site is introduced at the 5′-end;

[0039] P3 (SEQ ID NO.4): 5-GTCTCTAGAATCTGGAC...

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Abstract

The invention discloses a universal transfer vector for a pseudorabies virus capable of independently expressing dual genes and a construction method of the universal transfer vector. The universal transfer vector is obtained by inserting a dual-gene expression cassette containing two CMV promoters, multiple cloning sites respectively at downstream positions of the two CMV promoters and a BGH pA signal sequence between gEL and gER contained in pUC19 / HEK-gE recombinant plasmids containing gEL and gER. The universal transfer vector is capable of independently expressing two exogenous genes by utilizing two promoters and can be used for constructing a recombinant pseudorabies virus capable of independently expressing two exogenous genes.

Description

technical field [0001] The invention relates to the technical field of genetic engineering of animal viruses, in particular to a universal transfer vector of pseudorabies virus independently expressing two genes, its construction method and application. Background technique [0002] Rabies is an acute infectious disease caused by pseudorabies virus (PRV) infecting pigs, cattle, sheep, dogs, cats, rabbits, mice, mink, foxes and other domestic and wild animals. store and disseminator. Other animals except pigs are in sporadic form, and usually have symptoms such as fever, itching, and encephalomyelitis after the onset, all of which are fatal infections. The disease is an outbreak in pigs, mainly causing reproductive failure in sows and mass mortality in newborn piglets. PRV usually infects animals through the nasopharynx, proliferates in the infected local mucosa, and then migrates through the lymphatic circulation or nerve fibers. latent infection. [0003] The genome of ...

Claims

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Application Information

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IPC IPC(8): C12N15/63C12N5/10C12N15/66
Inventor 黄红亮陈瑞爱何玲谢小雨任常宝
Owner GUANGDONG WENS DAHUANONG BIOTECH
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