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Use of staphylococcal enterotoxin A gene and its coded protein

A technology of staphylococcal intestinal and toxin, applied in gene therapy, antibacterial, antiviral, etc., can solve problems such as weak immunogenicity

Inactive Publication Date: 2006-11-01
INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, some antigenic genes are less immunogenic and injection of these vaccines alone does not induce sufficient protective immunity

Method used

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  • Use of staphylococcal enterotoxin A gene and its coded protein
  • Use of staphylococcal enterotoxin A gene and its coded protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0012] Embodiment 1, the preparation of plasmid

[0013] 1. Construction of recombinant plasmid pmSEA containing staphylococcal enterotoxin A gene

[0014] Use upstream primer P1: 5'-cgc ggatcc agcgagaaaagcgaagaaa-3' (underlined base is the recognition site of restriction endonuclease BamH I) and downstream primer P2: 5'-atgaattca gaattc acttgtatataatatAGCaata tgcat-3' (the underlined base is the restriction endonuclease EcoR I recognition site), with the genomic DNA of Staphylococcus aureus standard strain ATCC 13565 as template, PCR amplification of SEA gene, the reaction conditions are: (1 ) 96°C for 10 minutes; (2) 94°C for 60 seconds, 50°C for 90 seconds, 72°C for 90 seconds, a total of 30 cycles; (3) 72°C for 10 minutes. After the reaction, the PCR product was subjected to agarose gel electrophoresis to recover a 702bp target fragment, and then the recovered gene fragment was digested with restriction endonucleases BamHI and EcoRI, and the digested product was cloned...

Embodiment 2

[0023] Embodiment 2, pmSEA enhances the immunogenicity of the hepatitis B virus surface antigen DNA vaccine

[0024] Mice: Balb / c, female, 4-6 weeks old.

[0025] Grouping: (1) Empty vector group (pcDNA3), 10 rats; (2) Hepatitis B protein vaccine group (rHBsAg), 10 rats; (3) pS2S group, 10 rats; (4) pS2S+pmSEA adjuvant group, 10 rats .

[0026] Immunization scheme: (1) Empty vector group (pcDNA3): 200ug / monkey / time; (2) Hepatitis B protein vaccine group (rHBsAg):

[0027] 400ng / animal / time; (3) pS2S group: 100ug pS2S+100ug pcDNA3 / animal / time; (4) pS2S+pmSEA adjuvant group: 100ug pS2S+100ug pmSEA / animal / time.

[0028] Under mouse anesthesia (75mg / kg sodium pentobarbital IP), 50uL samples were injected into the quadriceps muscles of the two hind limbs of the mouse, and then the in vivo gene transfer instrument (Zhejiang Xinzhi Company) was used to shock the injection site. The voltage is 120V / cm, the duration is 2ms, 2 times.

[0029] Number of immunizations: two times in to...

Embodiment 3

[0031] Example 3, pmSEA enhances the immunogenicity of malaria multi-epitope antigen DNA vaccine

[0032] Mice: Balb / c, female, 4-6 weeks old.

[0033] Grouping: (1) Empty vector group (pcDNA3), 6 rats; (2) AWTE plasmid group, 6 rats; (3) AWTE plasmid+pmSEA plasmid adjuvant group, 6 rats.

[0034] Immunization scheme: (1) Empty vector group (pcDNA3): 200ug / piece / time; (2) AWTE plasmid group: 100ugAWTE+100ug pcDNA3 / piece / time; (3) AWTE plasmid+pmSEA plasmid adjuvant group: 100ug AWTE+ 100ugpmSEA / piece / time.

[0035] Under mouse anesthesia (75mg / kg sodium pentobarbital IP), inject 50ul samples into the quadriceps muscles of the two hind limbs of the mouse respectively, and then use an in vivo gene transfer instrument (Zhejiang Xinzhi Company) to perform electric shock at the injection site, The voltage is 120V / cm, the duration is 2ms, 2 times.

[0036] Number of immunizations: two times in total. After the initial immunization, a booster immunization will be given on the 14th...

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Abstract

An application of staphylococcal enterotoxin A gene and its coding protein in obviously improving the responsiveness of animal to DNA vaccine and intensifying the immunogenicity of various DNA vaccines for hepatitis B virus, malaria, etc is disclosed.

Description

technical field [0001] The present invention relates to a new application of the staphylococcal enterotoxin A gene and its encoded protein, in particular to the use of the staphylococcal enterotoxin A gene as an adjuvant for enhancing the immunogenicity of DNA vaccines and the encoded protein of the gene as a recombinant subunit for enhancing the immunogenicity Use in adjuvants. Background technique [0002] Staphylococcal enterotoxin A (staphylococcal enterotoxin A, SEA) is a member of the staphylococcal enterotoxin family produced by Staphylococcus aureus and is one of the most effective T cell activating factors. SEA is a protein molecule containing 233 amino acid residues, rich in serine (S), threonine (T) and aspartic acid (D) residues. Small molecular weight, about 27.8kD, isoelectric point PI = 7.3, stable to acid and heat. There is a ring formed by disulfide bonds in the center of the SEA molecule, and the integrity of the ring is necessary for SEA to activate T ly...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61P1/16A61P31/04A61P31/12A61P33/00
CPCY02A50/30
Inventor 马清钧靳彦文胥全彬曹诚
Owner INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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