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Specific chicken CpG ODN and its application

A specific, DNA vaccine technology, applied in the field of CpG DNA, can solve the problems of weak immunogenicity and strong immune response of DNA vaccines, and achieve the effect of enhancing the protection rate of virus attack, increasing the antibody level, and enhancing the immune effect

Active Publication Date: 2006-08-02
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Many studies have shown that although the DNA vaccine plasmids of many H5 subtype AIV HA genes that have been constructed so far can produce certain immune protection responses, there are still weak immunogenicity of DNA vaccines, which are not as strong as the immune response caused by oil emulsion inactivated vaccines, etc. Insufficient, so further optimization is needed to enhance its immune efficacy

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0012] The preparation of embodiment 1CpG single-stranded deoxynucleotide

[0013] The ODN sequence was synthesized by Shanghai Yingjun Biotechnology Co., Ltd., using the ABI39X series synthesizer, using the solid-phase phosphoramidite triester method.

Embodiment 2

[0014] Preparation of the H5 subtype avian influenza DNA vaccine of embodiment 2 chimeric CpG

[0015] 1. PCR amplification of the HA cDNA coding region of H5 subtype avian influenza virus and the internal ribosome binding site (IRES) fragment of encephalomyocarditis virus (ECMV)

[0016] 1. Design and synthesis of primers

[0017] The primers used to amplify the HA gene cDNA of H5 subtype avian influenza virus and the internal ribosome binding site (IRES) fragment of encephalomyocarditis virus (ECMV) are designed with reference to the published sequence and aided by primer design software Primer Premier 5 made. A restriction site and a protective base are added to the 5' end of the primer. Primers were synthesized by Shanghai Boya Biotechnology Co., Ltd. The primer sequences are as follows:

[0018] HA P1: TA CTCGAG AAA ATG GAGAAAATAGTGCTT (XhoI)

[0019] HA P2: AT GGGCCC TACAATCTGAACTCACA (ApaI)

[0020] IRES P1: TC GGATCC TGCATCTAGGGCGGCCAA (BamHI)

[0021] IR...

Embodiment 3

[0047] The immunization application of the H5 subtype avian influenza DNA vaccine of embodiment 3 chimeric CpG

[0048] Recombinant Salmonella: ST (pVAX1-HA-CpG) and ST (pVAX1-HA) were first immunized with 3-day-old commercial Isha brown layer hens through oral and nasal routes, and the dose was 5×10 9 CFU / only; and the avian influenza oil emulsion inactivated vaccine immunization group, the empty vector ST (pVAX1) immunization group and the blank group were used as positive and negative controls, respectively. Booster immunization with the same immunization dose and immunization route two weeks after the first immunization, and challenge virus 3 weeks after the second immunization to evaluate the adjuvant effect of chimeric CpG DNA in the attenuated Salmonella ST delivery H5 subtype avian influenza nucleic acid vaccine, see Table 1.

[0049] serial number

[0050] **: p<0.01, the difference is extremely significant.

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Abstract

The present invention relates to CpG DNA, and is especially specific chicken CpG ODN and its use as the H5-subtype bird flu DNA vaccine adjuvant. The CpG ODN sequence is 5í»-TTGGAACGTTCCTTTCCAACGTTGGTTTGGAACGTTCCTT-3í». The H5-subtype bird flu DNA vaccine with the CpG ODN adjuvant is taken by chicken through oral taking or nose dropping to immunize, and can raise the protection rate against the challenge virus and the intestinal mucous membrane IgA antibody level of chicken obviously. The DNA vaccine of the present invention has the advantages of safety, no interference on the monitoring of bird flu epidemic situation, capacity of overcoming the interference of maternal antibody, use convenience, etc.

Description

technical field [0001] The present invention relates to a kind of CpG DNA (CpG DNA refers to the DNA that contains cytosine and guanine dinucleotide motif), particularly relates to the CpG ODN (ODN refers to oligonucleotide single-stranded DNA) that is used as chicken specificity ), and the use of this CpG ODN as an adjuvant for the H5 subtype avian influenza DNA vaccine. Background technique [0002] The control of highly pathogenic avian influenza (HPAI) mainly adopts isolation and blockade of epidemic sites, culling and destruction of infected and threatened animals, and other biosecurity measures. In view of the constraints of economic compensation pressure and other aspects, vaccination has also become one of the key measures to control the epidemic. The currently used oil emulsion whole virus inactivated vaccine has the advantages of definite effect, high protection rate, long immunity period and short development cycle. However, there are many defects in the use of ...

Claims

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Application Information

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IPC IPC(8): C07H21/04A61K48/00A61P31/16
Inventor 焦新安胡青海潘志明黄金林孙林殷月兰唐丽华张成全
Owner YANGZHOU UNIV
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