[Objective] The aim was to investigate the expression level of recombinant gene P1-2A3C of FMDV Asia I in different Bombyx mori varieties,so as to screen out the B.mori varieties suitable for the foreign gene expressi...[Objective] The aim was to investigate the expression level of recombinant gene P1-2A3C of FMDV Asia I in different Bombyx mori varieties,so as to screen out the B.mori varieties suitable for the foreign gene expression.[Method] The recombinant B.mori baculovirus rBmNPV（P1-2A3C） are injected into pupae of original species and hybrids of B.mori respectively.Then,the expression of antigen is detected by ELISA method and compared.[Result] The expression level of P1-2A3C significantly differed in different B.mori varieties,in which,the hybrids of Qiufeng×TQ78 and Qiufeng×Sijiaojian could be considered as the effective bioreactors for high-level expression of foreign genes.[Conclusion] This study provided the basis for breeding special B.mori varieties which can highly express target protein of Asia I FMDV.展开更多
[Objective] The aim was to study the prokaryotic expression of P1 gene of foot-and-mouth disease virus（FMDV）type Asia 1and the preparation of its antiserum.[Method]The P1 gene of FMDV type Asia 1 was obtained by gen...[Objective] The aim was to study the prokaryotic expression of P1 gene of foot-and-mouth disease virus（FMDV）type Asia 1and the preparation of its antiserum.[Method]The P1 gene of FMDV type Asia 1 was obtained by gene cloning techniques,and then cloned into pET-32a（＋）plasmid;subsequently the recombinant plasmid was transformed into E.coli BL21（DE3）;after the IPTG induction and protein purification,SDS-PAGE analysis was carried out;the ultrasonic wave was use to lyse the cultivated recombinant strain,and after the isolation and purification,this fusion protein was utilized to immunize New Zealand rabbits so as to prepare P1 protein antiserum.[Result]The positive clones were obtained;SDS-PAGE result showed that the target band was appeared at 105 kD;Western blot analysis showed that the antisera could bind to the expressed P1 fusion protein specifically;the ELISA titer of the rabbit anti-FMDV-P1 sera was approximately 1∶5 120.[Conclusion]This study had provided foundations for FMDV serological diagnostic methods and genetically engineered vaccine.展开更多
Monoclonal antibodies (McAbs) 1A9 and 9F12 against Foot-and-mouth disease virus (FMDV) serotype O were produced by fusing SP2/0 myeloma cells with splenocyte from the mouse immunized with O/China99. Both McAbs reacted...Monoclonal antibodies (McAbs) 1A9 and 9F12 against Foot-and-mouth disease virus (FMDV) serotype O were produced by fusing SP2/0 myeloma cells with splenocyte from the mouse immunized with O/China99. Both McAbs reacted with O/China99 but not with Asia 1, as determined by immunohistochemistry assay. The microneutralization titer of the McAbs 1A9 and 9F12 were 640 and 1 280, respectively. Both McAbs contain kappa light chains, but the McAbs 1A9 and 9F12 were IgG1 and IgM, respectively. In order to define the McAbs binding epitopes, the reactivity of these McAbs against VP1, P20 and P14 were examined using indirect ELISA, the result showed that both McAbs reacted with VP1 and P20. McAbs may be used for further studies of vaccine, diagnostic methods, prophylaxis, etiological and immunological researches on FMDV.展开更多
基金Supported by863Project for Development and Production of New FMDV accine for Livestock(2011AA10A211)Fund for Agricultural Since and Technology Achievements Transformation(2009GB23260437)~~
文摘[Objective] The aim was to investigate the expression level of recombinant gene P1-2A3C of FMDV Asia I in different Bombyx mori varieties,so as to screen out the B.mori varieties suitable for the foreign gene expression.[Method] The recombinant B.mori baculovirus rBmNPV（P1-2A3C） are injected into pupae of original species and hybrids of B.mori respectively.Then,the expression of antigen is detected by ELISA method and compared.[Result] The expression level of P1-2A3C significantly differed in different B.mori varieties,in which,the hybrids of Qiufeng×TQ78 and Qiufeng×Sijiaojian could be considered as the effective bioreactors for high-level expression of foreign genes.[Conclusion] This study provided the basis for breeding special B.mori varieties which can highly express target protein of Asia I FMDV.
基金Supported by National Transgenic Major Program of China(2009ZX08007-006B)the National Natural Science Foundation of China(31072160)+2 种基金Science and Technique Foundation of Shandong Province(2009GG20002032)Natural Science Foundation ofShandong Province(Y2008D20)an Open Issue of State Key Laboratory of Veterinary Biotechnology Fund(SKLVBF200806)~~
文摘[Objective] The aim was to study the prokaryotic expression of P1 gene of foot-and-mouth disease virus（FMDV）type Asia 1and the preparation of its antiserum.[Method]The P1 gene of FMDV type Asia 1 was obtained by gene cloning techniques,and then cloned into pET-32a（＋）plasmid;subsequently the recombinant plasmid was transformed into E.coli BL21（DE3）;after the IPTG induction and protein purification,SDS-PAGE analysis was carried out;the ultrasonic wave was use to lyse the cultivated recombinant strain,and after the isolation and purification,this fusion protein was utilized to immunize New Zealand rabbits so as to prepare P1 protein antiserum.[Result]The positive clones were obtained;SDS-PAGE result showed that the target band was appeared at 105 kD;Western blot analysis showed that the antisera could bind to the expressed P1 fusion protein specifically;the ELISA titer of the rabbit anti-FMDV-P1 sera was approximately 1∶5 120.[Conclusion]This study had provided foundations for FMDV serological diagnostic methods and genetically engineered vaccine.
基金The national high technology research and development program of China 863 (2006AA10A204)The national science and technology pillar program (2006BAD06A17)
文摘Monoclonal antibodies (McAbs) 1A9 and 9F12 against Foot-and-mouth disease virus (FMDV) serotype O were produced by fusing SP2/0 myeloma cells with splenocyte from the mouse immunized with O/China99. Both McAbs reacted with O/China99 but not with Asia 1, as determined by immunohistochemistry assay. The microneutralization titer of the McAbs 1A9 and 9F12 were 640 and 1 280, respectively. Both McAbs contain kappa light chains, but the McAbs 1A9 and 9F12 were IgG1 and IgM, respectively. In order to define the McAbs binding epitopes, the reactivity of these McAbs against VP1, P20 and P14 were examined using indirect ELISA, the result showed that both McAbs reacted with VP1 and P20. McAbs may be used for further studies of vaccine, diagnostic methods, prophylaxis, etiological and immunological researches on FMDV.
