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Recombinant salmonella SL7207 (pVAXD-N-Sa)

A technology of SL7207 and Salmonella, applied in the field of recombinant Salmonella SL7207, can solve the problems of poor immunogenicity and achieve the effect of good clinical application prospect, good safety and strong protection

Inactive Publication Date: 2015-06-17
SICHUAN AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

For example, the patent application whose publication number is CN 103194471A discloses a bigenic lactic acid bacteria vaccine comprising a segment of the porcine transmissible gastroenteritis virus S gene and an N gene, but the immunogenicity is not good

Method used

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  • Recombinant salmonella SL7207 (pVAXD-N-Sa)
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  • Recombinant salmonella SL7207 (pVAXD-N-Sa)

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Embodiment 1

[0060] Embodiment 1 Construction of recombinant bacteria of the present invention

[0061] 1. Preparation method

[0062] 1 Construction of double-promoter eukaryotic plasmid

[0063] 1.1 RT-PCR amplification of S gene and N gene

[0064] According to the S gene sequence of TGEV, a pair of primers were designed to amplify the 5’-1kb antigenic region of the S gene (named Sa, including four antigenic sites A, B, C, and D), and the upstream primer S1: 5’-CG GCTAGC ACCATGGAAAAACTATTTG-3', introduce restriction site NheI; downstream primer S2: 5'-CC AAGCTT TTAGCCACTAAGTAGCGTC-3', introduce restriction site HindIII. Design a pair of primers according to the N gene sequence, the expected amplification length is, upstream primer N1: 5'-CG GAATTC AAAATGGCCAACCAGGGAC-3', introduce restriction site EcoRI; downstream primer N2: 5'-CC CTCGAG AAGACGAGCATCTC-3', the restriction site XhoI was introduced.

[0065] Take TGEV cell culture virus liquid, extract viral RNA according to th...

experiment example 1

[0123] Experimental example 1 Characteristic identification of recombinant attenuated Salmonella of the present invention

[0124] 1. Experimental method

[0125] Get the recombinant Salmonella SL7207 (pVAX-S) prepared in Example 1 499-650 ), according to the following method for identification:

[0126] 1 Identification of biological characteristics of strain SL7207 (pVAXD-N-Sa)

[0127] 1.1 Microscopic examination of Gram staining of bacterial strain SL7207 (pVAXD-N-Sa) and control strain

[0128] SL7207(pVAXD-N-Sa) and control strains SL7207(pVAX-S), SL7207(pVAX-N), SL7207(pVAX-S-N) and SL7207(pVAXD) were respectively streaked on LB plates containing 100 μg / mL Kan, Cultivate for 16 hours, pick a single colony, inoculate 5 mL of LB liquid medium containing 100 μg / mL Kan, and culture at 37 ° C with shaking at 300 r / min until the OD600 is about 0.6, take the bacterial liquid for Gram staining and microscopic examination, observe and record the bacteria form.

[0129] 1.2 De...

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PUM

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Abstract

The invention discloses a nucleotide sequence represented by SEQ ID NO:1, a recombinant vector containing nucleotide sequences represented by SEQ ID NO:1 and SEQ ID NO:2 respectively, and a recombinant bacterium containing the nucleotide sequences. The recombinant salmonella SL7207 (pVAXD-N-Sa) containing the nucleotide sequences represented by SEQ ID NO:1 and SEQ ID NO:2 respectively is immune, can realize simultaneous inducible formation of transmissible two specific antibodies of the S protein and the N protein of gastro-enteritis virus (TGEV), has double-gene immune functions, comprehensive immune effects and high induced antibody level, can provide comprehensive immune protection effects for the transmissible gastro-enteritis, and has excellent clinic application prospect.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a recombinant Salmonella SL7207 (pVAXD-N-Sa). Background technique [0002] Porcine transmissible gastroenteritis (TGE) is a highly contagious intestinal infectious disease of pigs caused by porcine transmissible gastroenteritis virus (TGEV). , diarrhea and dehydration death, pigs of different breeds and ages are susceptible to the disease, especially the mortality rate of piglets within 2 weeks of age can be as high as 100%. Since 2010, intractable diarrhea in piglets has occurred in a large area of ​​pig herds in my country, and TGEV is one of the main pathogens frequently detected. [0003] At present, most of them are traditional inactivated vaccines and attenuated vaccines, but they have the risk of strong virulence and loose poison, which needs to be improved. In order to develop a porcine epidemic diarrhea vaccine that is safer, more efficient, less expensive, and easi...

Claims

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Application Information

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IPC IPC(8): C12N15/50C12N15/74C12N1/21C12R1/42
Inventor 黄小波文心田张鑫淼曹三杰张丹张雨迪文翼平伍锐马晓平张小会
Owner SICHUAN AGRI UNIV
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