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Novel use of thymosin alpha protogene

A technology of thymosin and pro-gene is applied in the application field of pro-thymosin alpha gene as an adjuvant for enhancing immunogenicity of DNA vaccine, which can solve the problems of weak immunogenicity and the like

Inactive Publication Date: 2006-07-26
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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  • Novel use of thymosin alpha protogene
  • Novel use of thymosin alpha protogene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0011] Example 1: Preparation of plasmid

[0012] 1. Construction of recombinant plasmid pThα containing prothymosin α gene

[0013] Use upstream primer 5’cgc ggatcc atgtctgatgcagctgtagatacc-3’ (BamHI), downstream primer 5’ccg gaattc gtc atcctcgtcggtcttctg-3'(EcoRI), using human fetal liver cDNA library as a template to amplify prothymosin α gene: (1) 95°C, 5 minutes; (2) 95°C, 30 seconds, 55°C, 20 seconds, 72°C, 30 seconds, 25 cycles; (3) 72°C, 5 minutes. The PCR product was recovered after gel electrophoresis, and the gene fragments obtained by digestion with KpnI and EcoRI were cloned into the plasmid vector pcDNA3 (Invitrogen, USA) that was digested with the same digestion to obtain the recombinant plasmid pThα. Transform DH5α Escherichia coli, select positive clones, and the DNA sequence analysis is correct (sequence 1), and the encoded amino acid sequence is sequence 2 in the sequence list. The restriction map of the plasmid is as follows figure 2 As shown in the figure, ...

Embodiment 2

[0022] Example 2: Enhancement of thymosin alpha gene on immunogenicity of hepatitis B virus surface antigen DNA vaccine

[0023] Mouse: C57BL / 6, female, 4-6 weeks old (19-21g).

[0024]Grouping: (1) empty vector group (pcDNA3), 6 animals; (2) hepatitis B protein vaccine group (rHBsAg), 6 animals; (3) pS2S group, 30 animals; (4) pS2S+pThα adjuvant group, 10 animals (5) TLH (fusion of hepatitis B surface antigen gene and prothymosin α gene) plasmid group, 6 mice.

[0025] Immunization program: (1) Empty vector group: pcDNA3200ug / head / time; (2) Hepatitis B protein vaccine group (rHBsAg): 400ng / head / time; (3) pS2S group: 100ug pS2S+100ug pcDNA3 / head / time; ( 4) pS2S+pThα adjuvant group: 100ug pS2S+100ug pThα. (5) TLH plasmid group: 100ug TLH.

[0026] Under the mouse anesthesia (75mg / kg sodium pentobarbital IP), 50ul of the sample was injected into the four femoral muscles of the two hind limbs of the mouse, and then the in vivo gene transfer instrument (produced by Zhejiang Xinzhi Com...

Embodiment 3

[0029] Example 3. Enhancement effect of prothymosin alpha gene on the immunogenicity of malaria multi-epitope antigen DNA vaccine

[0030] Mice: Balb / c, female, 4-6 weeks old (19-21g).

[0031] Grouping: (1) Empty vector group (pcDNA3), 6 animals; (2) AWTE plasmid group, 6 animals; (3) AWTE plasmid + prothymosin alpha (pThα) plasmid adjuvant group, 6 animals; (4) pTha -ATWE (malaria multi-epitope antigen gene fusion with prothymosin alpha gene) plasmid group, 6 mice.

[0032] Immunization protocol: (1) Empty vector group: pcDNA3 200ug / pcDNA3 / time; (2) AWTE plasmid group: 100ug AWTE+100ug pcDNA3 / pcDNA3 / time; (3) AWTE plasmid+thymosin α original plasmid adjuvant group: 100ug AWTE+100ugpThα. (4) pTha-ATWE plasmid group: 100ug pTha-ATWE.

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Abstract

The invention discloses an extrasin alpha original gene new usage, which is characterized by the following: the ProT alpha improves the reactivity of animal reacting to DNA vaccin observably, which can use as immune original-nature reinforcing adjuvant of DNA vaccin; the extrasin alpha original gene strengthens immune original-nature of hepatitis B virus surface antigen DNA vaccin, malarial disease multiple-epitope antigen DNA vaccin and kinds of DNA vaccines, which possesses broad-spectrum nature.

Description

Technical field [0001] The present invention relates to a new use of the prothymosin alpha gene, in particular to the application of the prothymosin alpha gene as an adjuvant for enhancing the immunogenicity of DNA vaccines. Background technique [0002] Prothymosin alpha (Prothymosin alpha, ProT alpha) is a small molecule protein composed of 109 to 113 amino acid residues. About 50% of the total number of amino acid residues in the ProTα molecule are acidic amino acid residues, which are distributed in clusters in the middle of the ProTα molecule, which makes the molecule highly hydrophilic. The ProTα molecule does not contain secretion signals, but contains a two-part nuclear localization signal, which is mainly distributed in the nucleus and is related to cell proliferation. The serum contains only a small amount of ProTα (about 10% of whole blood ProTα). Rat ProTα consists of 111 amino acid residues, and human ProTα consists of 110 amino acid residues. There are two types of ...

Claims

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Application Information

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IPC IPC(8): A61K39/39A61K48/00C12N15/09
CPCY02A50/30
Inventor 马清钧靳彦文曹诚
Owner INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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