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Method for expressing eimeria acevulina 3-1E protein in lactococcus lactis

A technology of Lactococcus lactis and Eimeria, applied in the field of expressing Eimeria gallinarum 3-1E protein, can solve the problem of inability to express Eimeria gallinarum sporozoite/merozoite stage surface antigen 3 -1E and other issues

Inactive Publication Date: 2012-12-12
NORTHEAST AGRICULTURAL UNIVERSITY
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Problems solved by technology

[0005] The present invention aims to solve the problem that Eimeria gallinarum sporozoite / merozoite stage surface antigen 3-1E cannot be expressed in Lactococcus lactis at present, and provides a method for expressing Eimeria gallinarum type 3 in Lactococcus lactis -1E protein method

Method used

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  • Method for expressing eimeria acevulina 3-1E protein in lactococcus lactis
  • Method for expressing eimeria acevulina 3-1E protein in lactococcus lactis
  • Method for expressing eimeria acevulina 3-1E protein in lactococcus lactis

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specific Embodiment approach 1

[0015] Specific embodiment one: In this embodiment, the method for expressing the Eimeria gallinacea 3-1E protein in Lactococcus lactis is carried out according to the following steps:

[0016] 1. Optimize the 3-1E gene sequence of Eimeria gallissima according to the codon preference of Lactococcus lactis, obtain the artificially synthesized 3-1E gene, and introduce BamH at its 5' end and 3' end respectively 2 and the recognition sequence of Xba 2, and then connected with the cloning vector pUC57 and transformed into Escherichia coli DH5α competent cells, and screened the positive transformant pUC57-3-1E;

[0017] 2. Digest pUC57-3-1E and Escherichia coli-lactic acid bacteria shuttle expression vector pTX8048 with BamH 3 and Xba3 respectively, recover the 3-1E fragment and vector fragment respectively, and construct the recombinant expression vector pTX8048 after ligation with T4 DNA ligase -3-1E;

[0018] 3. Transform pTX8048-3-1E into the competent cells of Lactococcus lact...

specific Embodiment approach 2

[0023] Specific embodiment 2: This embodiment is a further step of double enzyme digestion of pUC57-3-1E in step 2 of the method for expressing Eimeria gallinacea 3-1E protein in Lactococcus lactis described in specific embodiment 1 Instructions, the pUC57-3-1E double digestion system in step 2 is as follows:

[0024] Element

[0025] The enzyme digestion reaction condition was 37°C for 3h. Others are the same as in the first embodiment.

specific Embodiment approach 3

[0026] Specific embodiment three: this embodiment is a further description of the expression vector pTX8048 double enzyme digestion in step two of the method for expressing Eimeria gallinacea 3-1E protein in Lactococcus lactis described in specific embodiment one , the system of double digestion of the expression vector pTX8048 in step 2 is as follows:

[0027] Element

[0028] The enzyme digestion reaction condition was 37°C for 3h. Others are the same as in the first embodiment.

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Abstract

The invention relates to a method for expressing an eimeria acevulina 3-1E protein in lactococcus lactis, and relates to a method for expressing an eimeria acevulina 3-1E protein in order to solve the problem that the eimeria acevulina sporozoite / merozoite stage surface antigen 3-1E can not be expressed in the lactococcus lactis. The method comprises the following steps of optimizing an eimeria acevulina 3-1E gene sequence according to the preference of the condon of the lactococcus lactis, inserting a cloning vector and transforming escherichia coli cells to obtain positive transformants; carrying out enzyme digestion and connection of the positive transformants and a lactic acid bacteria expression vector, and constructing a recombinant expression vector; transforming competent lactic acid bacteria cells, and identifying; and culturing and inducing recombinant bacteria to complete the expression of the 3-1E protein in the lactococcus lactis NZ9000. The artificially synthesized codon-optimized 3-1E protein is successfully expressed in the lactococcus lactis. The method provided by the invention is used for the prevention field of coccidiosis.

Description

technical field [0001] The invention relates to a method for expressing Eimeria gallinacea 3-1E protein. Background technique [0002] Chicken Eimeria is one of the important diseases that seriously endanger the development of poultry industry. Chicken coccidia can cause epithelial cell damage and inflammation during the reproduction process in the body. Extensive damage can cause severe clinical diseases such as diarrhea, dehydration, weight loss, rectal prolapse, and dysentery in chickens, and sometimes death of chickens. The annual economic loss caused by coccidiosis in the world is about 3 billion U.S. dollars, 80% of which are due to the direct causes of chicken weight gain reduction, feed conversion rate reduction and death, and 20% are due to the cost of chemical drug research. It has been confirmed that frequent use of chemical drugs can lead to the emergence of drug-resistant strains, and drug residues can cause problems such as animal-derived food safety. The hig...

Claims

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Application Information

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IPC IPC(8): C12N15/74C12R1/01
Inventor 马德星马春丽冯兴军高铭扬葛俊伟李广兴李一经
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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