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Construction method and application of faeG expressing gene engineering probiotic

A technology of genetic engineering and construction method, which is applied in the application field of preventing Escherichia coli diarrhea in piglets, and achieves the effects of resisting degradation and destruction, making vaccines easy and improving immune effect.

Inactive Publication Date: 2011-04-27
ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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  • Construction method and application of faeG expressing gene engineering probiotic
  • Construction method and application of faeG expressing gene engineering probiotic
  • Construction method and application of faeG expressing gene engineering probiotic

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Embodiment Construction

[0022] The recombinant probiotics expressing enterotoxigenic Escherichia coli adhesin gene faeG described in the present invention are constructed by the following method: according to the published gene sequence and expression vector plasmid characteristics of F4 Escherichia coli pili adhesin FaeG, a specific The primers of the enzyme cutting site; using the enterotoxigenic Escherichia coli plasmid DNA isolated or purchased from diarrheal piglets as a template, PCR amplification was carried out to obtain a fragment containing the faeG gene, which was connected to the expression vector plasmid pNZ8148, Obtain the recombinant plasmid pNZ8148-faeG, transfer the recombinant plasmid into Lactobacillus or Lactococcus cells by electroporation, and express under the induction of nisin.

[0023] Among them, F4 Escherichia coli includes three serotypes of F4ab, F4ac and F4ad; the expression vector plasmid is pNZ8148 or the plasmid obtained after changing the selection marker based on pN...

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Abstract

The invention discloses a construction method and application of a faeG expressing gene engineering probiotic. The method comprises the following steps of: according to the gene sequence and the expression vector plasmid characteristic of a disclosed F4 escherichia coli pilus adhesin faeG, designing a primer which contains a specific endonuclease site; performing polymerase chain reaction (PCR) amplification by taking deoxyribonucleic acid (DNA) of an enterotoxigenic escherichia coli plasmid as a template to obtain a faeG gene-containing segment, linking the segment with an expression vector plasmid pNZ8148 to obtain a recombinant plasmid pNZ8148-faeG, transferring the recombinant plasmid into lactobacillus or a galactococcus cell by using an electrotransformation method to obtain gene engineering probiotic which expresses enterotoxigenic escherichia coli pilus adhesin faeG, expressing under the induction of nisin and analyzing and proving a target gene by SDS-PAGE and Western blot so as to express correctly. A recombinant strain can be used for preventing colibacillus diarrhea of suckling pigs and weanling pigs.

Description

technical field [0001] The invention belongs to the technical field of animal husbandry and veterinary medicine, and in particular relates to the construction of a genetically engineered probiotic bacterium expressing the enterotoxigenic Escherichia coli adhesin gene faeG and its application in preventing Escherichia coli diarrhea in piglets. Background technique [0002] Piglet diarrhea caused by Enterotoxigenic Escherichia coli (ETEC) poses a serious threat to pig production, and the pig industry suffers huge economic losses every year. Escherichia coli diarrhea in piglets includes diarrhea in suckling piglets and diarrhea in weaned piglets. Escherichia coli diarrhea in suckling piglets is divided into yellow scour and pullorum. Yellow dysentery of piglets is an acute and fatal infectious disease that often occurs in newborn piglets. It mainly occurs in suckling pigs within 7 days of age, and occurs frequently in suckling pigs of 1 to 3 days of age. After the onset, it is...

Claims

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Application Information

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IPC IPC(8): C12N15/31C12N15/63C12N1/21A61K39/108A61P1/12A61P31/04C12R1/46C12R1/225
Inventor 徐子伟刘淑杰李永明王一成
Owner ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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