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Preparation method and application of chlamydophila psittaci recombinant protein GST-CPSIT_p7

A technology of recombinant protein and chlamydia, applied in the field of genetic engineering, can solve the problems of unclear pathogenic mechanism of Cps, repeated infection, pathological damage, etc., to meet the needs of clinical diagnosis, improve sensitivity, and easy to operate

Inactive Publication Date: 2015-10-14
NANHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The intracellular parasitism of Cps often causes recessive and persistent infection, which makes the infection repeated and protracted, resulting in progressive and irreversible pathological damage
So far, the pathogenic mechanism of Cps is still unclear

Method used

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  • Preparation method and application of chlamydophila psittaci recombinant protein GST-CPSIT_p7
  • Preparation method and application of chlamydophila psittaci recombinant protein GST-CPSIT_p7
  • Preparation method and application of chlamydophila psittaci recombinant protein GST-CPSIT_p7

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] A preparation method for the recombinant protein GST-CPSIT_p7 of Chlamydia psittaci, comprising the following steps:

[0051] Step 1: Construct the recombinant expression plasmid pGEX-6P-1 / CPSIT_p7: Construct the recombinant expression plasmid pGEX-6P-1 / CPSIT_p7: Take the DNA of the Cps 6BC strain as a template and design the specific primers for CPSIT_p7:

[0052] Upstream primer, 5'-CGCGGATCCATGGGTAATTCTGGTTTTTACTT-3'; Downstream primer, 5'-TTTTCCTTTTGCGGCCGCTTAACCATTTGTTTGTTGTTTTAC-3';

[0053] After pre-denaturing the CPSIT_p7 region at 94°C for 5 minutes, it was denatured at 94°C for 30 seconds, annealed at 52°C for 45 seconds, and extended at 72°C for 120 seconds for 30 cycles. After the last cycle, it was extended at 72°C for 2 minutes to terminate the reaction and complete the PCR amplification. Increase, separate the polymerase chain reaction product through agarose electrophoresis, and purify the obtained PCR amplification product, comprising the following ste...

Embodiment 2

[0072] Example 2: Preparation and Performance Test of the Kit for Detecting Chlamydia psittaci Infection

[0073] The recombinant protein GST-CPSIT_p7 can be applied in the kit for detecting Chlamydia psittaci infection, as the antigen in the kit components for detecting Chlamydia psittaci infection, the antigen is labeled with colloidal gold, and the CT is measured by colloidal gold method Antibody. The preparation method of the kit that detects psittacosis chlamydia infection:

[0074] (1) Take 1.0g of HAuCl 4 4H 2 O is dissolved in 100ml purified water to make 1% chloroauric acid solution; get 1ml of 1% (mass concentration, mass concentration is not specified in this application) chloroauric acid solution into 100ml boiling water, add 2ml, 1% of trisodium citrate, and continue to boil for 30 minutes to synthesize colloidal gold; take 6ml of the synthesized colloidal gold solution, and scan it at 400-700nm; take 406ml of the synthesized colloidal gold of about 30nm 2 CO ...

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Abstract

The invention relates to the technical field of genetic engineering, in particular to a preparation method and application of chlamydophila psittaci recombinant protein GST-CPSIT_p7. The method comprises the following steps: step I: constructing recombinant expression plasmid pGEX-6P-1 / CPSIT_p7; step II: inducing the recombinant expression plasmid pGEX-6P-1 / CPSIT_p7 to express; and step III: preparing the recombinant protein GST-CPSIT_p7. By adopting the method, the chlamydophila psittaci recombinant protein GST-CPSIT_p7 is successfully cloned, expressed and purified, and the prepared protein has better immunogenicity. The recombinant protein GST-CPSIT_p7 can be applied to kits used for detecting the chlamydophila psittaci infection, and has the advantages of high specificity, high sensitivity and simplicity and convenience in operation, and clinical diagnosis needs of chlamydophila psittaci infection are met better.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to a preparation method and application of a Chlamydia psittaci recombinant protein GST-CPSIT_p7. Background technique [0002] Chlamydiaceae (Chlamydiaceae) are a class of obligate intracellular parasitic Gram-negative bacteria that can cause various diseases in humans and animals. With the in-depth research on Chlamydia, it was divided into Chlamydia and Chlamydophila according to 16S rRNA and 23S rRNA in 1999. The genus Chlamydia can be divided into three species: C. trachomatis, C. muridarum, and C. suis; the genus Chlamydia can be divided into C. psittaci, abortion Chlamydia (C.abortus), guinea pig Chlamydia (C.caviae), cat Chlamydia (C.felis), animal Chlamydia (C.pecorum), pneumoniae Chlamydia (C.pneumoniae) 6 species. The ones that have a greater impact on humans are Chlamydia trachomatis (Ct), Chlamydia pneumoniae (Cpn) and Chlamydia psittaci (Cps). Cps can ca...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/70C12N15/31C07K19/00G01N33/68
Inventor 贺庆芝曾怀才康颖杨超
Owner NANHUA UNIV
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