| Some molecule Biological characteristic of Egg Drop Syndrome Virus (EDSV) SG9301 strain that was isolate from Sichuan province in China was studied in this paper.1. Restriction endonucleases analysis. Using nine kinds restriction endonucleases (RE) (BamH I , Kpn I Hind III Pst I Sma I Bg1 I Bg1 II EcoR I ,EcoRV), DNA of EDSV-SG9301 strain was digested, the size of fragments were calculated and also compared with those of AV-127, H91, NE4, Qac94, JE1 .The results showed that the RE maps were similar each other, but were not same between any strains, and the comparability of the RE maps of EDSV strains was consistent with the origin of virus from animal and the similarity of symptom. The test indicated the RE analysis was a useful tool in differentiating EDSV strains and epidemical survey.2. Cloning of the hexon-encoding gene. According to the nucleotide sequence of EDSV-AV127 strain from the Genbank, a pairs of oligonucleotide primer was designed by using Oligo program and synthesized. With use of polymerase chain reaction (PCR), the hexon-encoding gene fragment was amplified from the genome of EDSV-SG9301 strain and the fragment directly inserted into pUCIS vector at EcoRl and Pstl site, the recombinant plasmid DNAs were used to transform the competent E. Coli JM109 cells. The positive colonies harboring the hexon-encoding gene were identified by digestion of REs, PCR, Southern hybrid and bolt hybrid .In the end, a positive clone named PUC-H3 was sequenced, and the result indicate that the plasmid contain the complete sequence of hexon-encoding gene.3. Sequencing of a part of the nucleotide and heredity-variance analysis. After analysis EDSV genome structure, hexon-encoding gene fragment and a nucleotide fragment that lie between PVI gene and fiber gene (from 22744 to 22367) was select to sequence and analyze. With use of PCR, the two fragments were amplified from the genome of EDSV SG9301 strain and the amplified fragments were inserted into pMD-T vector respectively. The recombinant plasmids were sequenced. The result showed that the hexon-encoding gene of EDSV SG9301 strain was 2733 base pair long which encoded 910 amino acids(aa). Comparison with AV-127 strain indicated thatthere was 99.85% homology at the nucleotide level, 99.78% homology at the amino acid level. Comparison with AAV-2 showed that there was 98.46% homology at the amino acid level. The results of nucleotide and amino acid comparison indicate that the biology character is consistent with the homologies of nucleotide and amino acid.The nucleotide sequence that lie between PV1 gene and fiber gene of EDSV SG9301 strain was compared with AV-127 , the result showed that the nucleotide sequence of the area was 99.85% homology and has not ORFs that the number of amino acid exceed 60aa .Analysis indicate that the hexon-encoding gene was quite conserved and the nucleotide sequence that lie between the PVi gene and fiber was more vary.4. Expression of the Hexon-Encoding Gene of EDSV-SG9301 Strain. A recominant plasmid named PUC-H3 that contains complete sequence of hexon-encoding gene was digested by restriction endonucleases (Pstl/EcoRl) , the hexon-encoding gene was cloned into the expression vector pBV220, one recominant plasmid named PPE24 was constructed which highly expressed a HOKDa protein in E.coli JM109. The result of SDS-PAGE, Western blotting and agar diffusion test indicate the expressed product could react with EDSV positive serum.Also, the hexon-encoding gene was cloned into the expression vector pQE32. A recominant plasmid named PPHE was constructed which highly expressed a HOKDa protein with 6Xhis in E.coli M15(PREP4) The result of SDS-PAGE, Western blotting and agar diffusion test indicate the expressed product could react with EDSV positive serum.5. Establishing detection means of biotinylated nucleic acid probe After labelling the recombinant plasmid harboring the hexon-encoding gene with biotin, the biotinylated recombinant plasmid DNA was as probe to detect EDSV-DNA. The tes...
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