[Research on secretion expression in Pichia pastoris and function of the HC-pro gene of watermelon mosaic virus].
HC-pro gene of Watermelon Mosaic virus was obtained by RT-PCR was 1371bp in length. It was cloned into pPI(9K, then the eucaryotic recombinant expression plasmid pPIC9K-WHC was constructed. After being linearized with restriction endonuclease Sal I , the recombinant plasmid was transformed into Pichia pastoris GS115 by electroporation. The high copy transformants with Mut+ /His+ phenotype were selected by RT-PCR and screening on G418, MD and MM medium. Induced by methanol for 5 days, the culture supernatant was analyzed by SDS-PAGE, the results showed that a specific protein with a molecular weight of about 66 kD was expressed. Western blot analysis proved that the expression protein could specifically bind to HC-Pro polyclonal antibody. Far western blot analysis proved that the expression protein could bind to coat protein, given support to "bridge" hypothesis that HC-Pro help aphid transmission of non-persistent viruses.
Zhang
Jian-Xin
JX
Colllege of Plant Protection and Shaanxi Key Laboratory of Molecular Biology for Agriculture, Northwest A&F University, Yangling 712100, China.
Wu
Yun-Feng
YF
Wang
Xiu-Min
XM
D029773:Citrullus / Q000821:virology*; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000235:genetics*; D009029:Mosaic Viruses / Q000235:genetics*; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000096:biosynthesis / Q000235:genetics*
D011994:Recombinant Proteins; D014764:Viral Proteins; D003546:Cysteine Endopeptidases; C072775:HC-Pro protein, potyvirus