[ Objective] The study aimed to understand the genetic characters of H9N2 subtype avian influenza viruses isolated in Belling area. [ Method] HA genes of three H9N2 subtype avian influenza viruses A/Chicken/Beijing/xu...[ Objective] The study aimed to understand the genetic characters of H9N2 subtype avian influenza viruses isolated in Belling area. [ Method] HA genes of three H9N2 subtype avian influenza viruses A/Chicken/Beijing/xu/00, A/Chicken/Beijing/bei/00 and A/Chicken/Beijing/ liu/00 were amplified by RT-PCR and then sequenced. [ Result] The results of phylogenetic analysis showed that A/Chicken/Beijing/xu/00, A/ Chicken/Beijing/bei/00 and A/Chicken/Beijing/liu/00 shared the nucleotide homologies of 84.8% （ Dk/HK/Y439/97 ） -98.0% （ Ck/GX17/00 ）, 85.1% （Dk/HK/Y439/97） - 99.1% （ Ck/GXl 7/00）, 90.7% （ Ck/BJ/3/01 ） - 99.1% （Ck/GX17/00） with the isolates from Hongkong and other are- as of Chinese Mainland respectively. At the same time, the analysis of amino acid indicated that the three isolates belonged to low pathogenic H9N2 isolates of avian origin. The 226^th amino acid of them were L （ Leu）, suggesting their high binding affinity to human cells. There were seven glyco- sylation sites in HA protein, five from HA1 and two from HA2. [ Cenclusien] By analysis at molecular level, it could be concluded that A/Chicken/ Beijing/xu/00, A/Chicken/Beijing/bei/00 and A/Chicken/Beijing/liu/00 were low pathogenic H9N2 isolates of avian origin.展开更多
HA Gene of H9N2. sub-type avian influenza virus from three strains in different times was amplified, purified and then sequenced. The results of its sequence analysis showed that the whole length of the amplified HA G...HA Gene of H9N2. sub-type avian influenza virus from three strains in different times was amplified, purified and then sequenced. The results of its sequence analysis showed that the whole length of the amplified HA Gene was 1 683 bp, encoding 560 amino acids. The amino acid sequence of three virulent strains at cleavage site was R-S-S-R, which was low-pathogenicity strain. According to the amino acid sequence of the isolated strains, there were 7 potential glycosylation sites, and the receptor-binding site was the specific sequence of the avian-derived influenza virus. Amino acids on the left edge of receptor-binding site were all NGQQG, while amino acids on the right edge of receptor-binding site were GTSKA. From the comparative sequence analysis of HA Gene from some referenced strains, the results indicated that nucleotide and amino acid homology between isolated strains and referenced strains was higher. Evolutionary tree analysis showed that three strains were all Eurasian species, and there was a close relationship with the representative strains of A / duck / Hong Kong/Y280/97.展开更多
To construct a recombinant adenovirus shuttle plasmid pDC315-H5HA-EGFP,the HA gene of A/Swine/Fujian/1/2001(H5N1) was amplified by RT-PCR and then inserted into adenovirus shuttle plasmid pDC315.A replication-defectiv...To construct a recombinant adenovirus shuttle plasmid pDC315-H5HA-EGFP,the HA gene of A/Swine/Fujian/1/2001(H5N1) was amplified by RT-PCR and then inserted into adenovirus shuttle plasmid pDC315.A replication-defective recombinant adenovirus expressing the HA gene(rAd-H5HA-EGFP) was generated by co-transfecting the recombinant shuttle plasmid pDC315-H5HA-EGFP and the genomic plasmid pBHGlox△E1,E3Cre in HEK293 cells.The recombinant adenovirus was confirmed by PCR,RT-PCR and Western blot assay.These results demonstrated that HA protein was properly expressed by the rAd-H5HA-EGFP in HEK293 cells and had natural biological activities.The TCID<sub>50</sub> of the rAd-H5HA- EGFP was assessed to be 2.26×10<sup>10</sup>/mL after propagation and purification.Immunization of BALB/ c mice indicated that rAd-H5HA-EGFP induced HI antibodies and protected mice from replication of the challenge virus in their lungs.展开更多
A neural network classification method,and a batch-learning self-organizing map(BLSOM),was established using trinucleotide and tetranucleotide in the hemagglutinin gene sequences of 25 avian influenza viruses isolated...A neural network classification method,and a batch-learning self-organizing map(BLSOM),was established using trinucleotide and tetranucleotide in the hemagglutinin gene sequences of 25 avian influenza viruses isolated in Guangdong Province. Statistical analysis and normalization of the fragment number were done and MATLAB function was used to simulate the human brain thinking for self-organizing learning. When the number of training steps was 100 and above,the strains could be successfully clustered. H_1,H_3,H_5,H_7 and H_9 subtype strains fell within different classes,respectively,and the HA gene cluster map of H_3N_2 and H_7N_9strains was quite similar,suggesting that these strains shared the same origin; H_5N_1 strain was quite different in different years; H_1N_1 and H_9N_2 strains could be clustered into one group,indicating the natural recombinant variation in the two kinds of viruses,thereby providing a reference for high-risk strain screening and traceability.展开更多
[Objective] The paper was to study the possible mechanism of chitinase gene. [Method] The chitinase gene with inhibitory effect against a variety of pathogenic fungi was cloned and analyzed in this study. [Result] The...[Objective] The paper was to study the possible mechanism of chitinase gene. [Method] The chitinase gene with inhibitory effect against a variety of pathogenic fungi was cloned and analyzed in this study. [Result] The genome of HAS strain contained an ORF of 834 bp. The homology of coding protein corresponding to ORF with chitosanase [Bacillus subtilis subsp. subtilis str. 168](ref|NP_390566.1) was the highest of 98%(272/277).[Conclusion]Through the analysis of amino acid sequence and secondary structure of the protease, tertiary structure prediction and analysis, HAS strain may play a pivotal role in controlling Sporisorium scitaminea Syd.展开更多
文摘[ Objective] The study aimed to understand the genetic characters of H9N2 subtype avian influenza viruses isolated in Belling area. [ Method] HA genes of three H9N2 subtype avian influenza viruses A/Chicken/Beijing/xu/00, A/Chicken/Beijing/bei/00 and A/Chicken/Beijing/ liu/00 were amplified by RT-PCR and then sequenced. [ Result] The results of phylogenetic analysis showed that A/Chicken/Beijing/xu/00, A/ Chicken/Beijing/bei/00 and A/Chicken/Beijing/liu/00 shared the nucleotide homologies of 84.8% （ Dk/HK/Y439/97 ） -98.0% （ Ck/GX17/00 ）, 85.1% （Dk/HK/Y439/97） - 99.1% （ Ck/GXl 7/00）, 90.7% （ Ck/BJ/3/01 ） - 99.1% （Ck/GX17/00） with the isolates from Hongkong and other are- as of Chinese Mainland respectively. At the same time, the analysis of amino acid indicated that the three isolates belonged to low pathogenic H9N2 isolates of avian origin. The 226^th amino acid of them were L （ Leu）, suggesting their high binding affinity to human cells. There were seven glyco- sylation sites in HA protein, five from HA1 and two from HA2. [ Cenclusien] By analysis at molecular level, it could be concluded that A/Chicken/ Beijing/xu/00, A/Chicken/Beijing/bei/00 and A/Chicken/Beijing/liu/00 were low pathogenic H9N2 isolates of avian origin.
文摘HA Gene of H9N2. sub-type avian influenza virus from three strains in different times was amplified, purified and then sequenced. The results of its sequence analysis showed that the whole length of the amplified HA Gene was 1 683 bp, encoding 560 amino acids. The amino acid sequence of three virulent strains at cleavage site was R-S-S-R, which was low-pathogenicity strain. According to the amino acid sequence of the isolated strains, there were 7 potential glycosylation sites, and the receptor-binding site was the specific sequence of the avian-derived influenza virus. Amino acids on the left edge of receptor-binding site were all NGQQG, while amino acids on the right edge of receptor-binding site were GTSKA. From the comparative sequence analysis of HA Gene from some referenced strains, the results indicated that nucleotide and amino acid homology between isolated strains and referenced strains was higher. Evolutionary tree analysis showed that three strains were all Eurasian species, and there was a close relationship with the representative strains of A / duck / Hong Kong/Y280/97.
基金supported by the Chinese National S&T Plan(2004BA519A55)Scientific Research Program of State Key Laboratory of Veterinary Biotechnology(NKLVBP200818)
文摘To construct a recombinant adenovirus shuttle plasmid pDC315-H5HA-EGFP,the HA gene of A/Swine/Fujian/1/2001(H5N1) was amplified by RT-PCR and then inserted into adenovirus shuttle plasmid pDC315.A replication-defective recombinant adenovirus expressing the HA gene(rAd-H5HA-EGFP) was generated by co-transfecting the recombinant shuttle plasmid pDC315-H5HA-EGFP and the genomic plasmid pBHGlox△E1,E3Cre in HEK293 cells.The recombinant adenovirus was confirmed by PCR,RT-PCR and Western blot assay.These results demonstrated that HA protein was properly expressed by the rAd-H5HA-EGFP in HEK293 cells and had natural biological activities.The TCID<sub>50</sub> of the rAd-H5HA- EGFP was assessed to be 2.26×10<sup>10</sup>/mL after propagation and purification.Immunization of BALB/ c mice indicated that rAd-H5HA-EGFP induced HI antibodies and protected mice from replication of the challenge virus in their lungs.
基金Supported by National Entry-Exit Inspection and Quarantine Research Project(2015IK054)Special Project of Ministry of Science and Technology for Development of Major Scientific Instruments and Equipments(2012YQ0901-9705)
文摘A neural network classification method,and a batch-learning self-organizing map(BLSOM),was established using trinucleotide and tetranucleotide in the hemagglutinin gene sequences of 25 avian influenza viruses isolated in Guangdong Province. Statistical analysis and normalization of the fragment number were done and MATLAB function was used to simulate the human brain thinking for self-organizing learning. When the number of training steps was 100 and above,the strains could be successfully clustered. H_1,H_3,H_5,H_7 and H_9 subtype strains fell within different classes,respectively,and the HA gene cluster map of H_3N_2 and H_7N_9strains was quite similar,suggesting that these strains shared the same origin; H_5N_1 strain was quite different in different years; H_1N_1 and H_9N_2 strains could be clustered into one group,indicating the natural recombinant variation in the two kinds of viruses,thereby providing a reference for high-risk strain screening and traceability.
基金Supported by Natural Science Foundation of China(31471555)National Key Research and Development Program "Integrated Research and Demonstration of Fertilizer and Pesticide Reduction Technology for Featured Cash Crops"(2018YFD0201100)
文摘[Objective] The paper was to study the possible mechanism of chitinase gene. [Method] The chitinase gene with inhibitory effect against a variety of pathogenic fungi was cloned and analyzed in this study. [Result] The genome of HAS strain contained an ORF of 834 bp. The homology of coding protein corresponding to ORF with chitosanase [Bacillus subtilis subsp. subtilis str. 168](ref|NP_390566.1) was the highest of 98%(272/277).[Conclusion]Through the analysis of amino acid sequence and secondary structure of the protease, tertiary structure prediction and analysis, HAS strain may play a pivotal role in controlling Sporisorium scitaminea Syd.
