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Chinese Journal of Biotechnology ; (12): 1068-1073, 2010.
Article in Chinese | WPRIM | ID: wpr-292170


On the basis of successful cloning the full length hemagglulinin (HA) and neuramidinase (NA) gene and sequence analysis of influenza virus H1N1, part of the gene was ligated into pMETA. Expression vectors pMETA/HA (52-1 557 bp) and pMETA/NA (121-1 263 bp) were constructed and expressed in pMAD16 induced by methanol. Recombinant protein was purified through Ni2+ affinity chromatography. Western blotting and ELISA were used to determine the antigenic activity of the recombinant protein. SDS-PAGE showed that the recombinant capsid gene could be overexpressed in Pichia methanolica. ELISA and Western blotting showed that the recombinant protein had antigenicity.

Cloning, Molecular , Genetic Vectors , Genetics , Hemagglutinins , Genetics , Influenza A Virus, H1N1 Subtype , Genetics , Neuraminidase , Genetics , Pichia , Genetics , Metabolism , Recombinant Proteins , Genetics , Allergy and Immunology
Progress in Biochemistry and Biophysics ; (12): 685-690, 2006.
Article in Chinese | WPRIM | ID: wpr-408521


A new and effective method to produce transgenic animals was established. Without a surgical incision, the recombinant plasmid containing green fluorescence protein (GFP) cDNA was repeatedly injected into male mouse testis at multi-sites. After few weeks of the final injection, the injected male was mated with normal oestrus female to produce transgenic mice. The presence of the GFP cDNA in F1 transgenic individuals were detected by polymerase chain reaction and Southern blot hybridization, which showed that the transgenic rate of mouse F1 offspring was 41%. The transferred gene was integrated into the host genome and could be transmitted to its offspring. When the positive F1 individuals were mated with the wild type ICR mice, the F2 individuals had a transgenic rate of 37%. The results indicate that the high efficiency of gene transfer and the limited number of manipulations make the method suitable for creating a large number of transgenic animals, especially, for producing domestic animals.