Preparation method for polyclonal antibody to salmonella effect protein SopB

A polyclonal antibody and effector protein technology, applied in the biological field, can solve problems such as no research reports, and achieve the effect of avoiding non-specific cleavage

Inactive Publication Date: 2014-06-18
TIANJIN UNIV OF COMMERCE
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  • Abstract
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Problems solved by technology

After searching the literature, it is found that there is no research report related to the preparation of Salmonella effector protein SopB antibody

Method used

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  • Preparation method for polyclonal antibody to salmonella effect protein SopB
  • Preparation method for polyclonal antibody to salmonella effect protein SopB
  • Preparation method for polyclonal antibody to salmonella effect protein SopB

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Embodiment

[0045] biomaterials

[0046] Salmonella enterica subsp.enterica serovar Typhimurium LT2 (Salmonella enterica subsp.enterica serovar Typhimurium) is deposited in the General Microbiology Center (CGMCC) of the China Committee for the Collection of Microbial Cultures. , the date of deposit is December 20, 2012, and the deposit number is CGMCC No.7020.

[0047] The pET28a expression vector was purchased from Novagen; Escherichia coli DH5α and Escherichia coli BL21(DE3) strains were purchased from Promega; restriction enzymes NdeI, XhoI, BamHI, T4 DNA ligase, Taq DNA polymerase and bacterial genomic DNA extraction kit All were purchased from Takara Company; pSUMO expression vector, pGEX-6P-2 expression vector, HiTrap Q HP anion exchange column, Superdex20010 / 300 molecular sieve and HiTrap NHS-activated HP affinity chromatography column were purchased from GE Healthcare; trishydroxymethylamino Methane, acrylamide, kanamycin, ampicillin, methylene bisacrylamide, sodium chloride, imi...

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Abstract

The invention discloses a preparation method for a polyclonal antibody to salmonella effect protein SopB. The invention aims to provide the preparation method for the rabbit-derived polyclonal antibody with good specificity, high sensitivity and capability of specifically binding to the endogenous salmonella effect protein SopB secreted in the process of salmonella infection of a host cell. A protein sequence as represented by SEQ ID No. 2 is used as immunogen, wherein the protein sequence consists of 139 amino acids located between position 29 and position 168, counted from terminal N, of a full-length amino acid sequence as represented by SEQ ID No. 1 of the salmonella effect protein SopB, and a conventional preparation method for a polyclonal antibody is used for preparation of antiserum; then the antiserum is purified so as to obtain the polyclonal antibody to the salmonella effect protein SopB. According to the invention, the polyclonal antibody to the salmonella effect protein SopB is prepared by using the method, and the polyclonal antibody is beneficial for immunoblotting analysis and immunofluorescent staining laser confocal microscopic observation of the salmonella effect protein SopB and for in-depth research on the infection mechanism of enteropathogens.

Description

technical field [0001] The invention relates to an antibody in the field of biotechnology and a preparation method thereof, in particular to a preparation method of a polyclonal antibody of Salmonella effector protein SopB. Background technique [0002] The Salmonella effector protein SopB is a protein molecule composed of 561 amino acids secreted by the Salmonella SPI-1 type III secretion system. It is a kind of inositol phospholipase. an egg white. The Salmonella effector protein SopB first acts on the plasma membrane, activates the SH3-guanylate exchange factor (SGEF), and indirectly activates the exchange factor RhoG, thereby further activating actin, inducing cytoskeleton rearrangement, and promoting Salmonella to invade host cells. The Salmonella effector protein SopB activates the proto-oncogene Akt 473 Phosphorylation of Salmonella inhibits cell apoptosis mediated by bacterial invasion, promotes the survival of host cells, and provides conditions for the survival o...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/40C07K16/06C12N15/55C12N15/70C12N9/16
Inventor 阮海华张振奇陶永清赵辉张坤生
Owner TIANJIN UNIV OF COMMERCE
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