tailieunhanh - Báo cáo khoa học: Production of a recombinant mouse monoclonal antibody in transgenic silkworm cocoons

In the present study, we describe the production of transgenic silkworms expressing a recombinant mouse mAb in their cocoons. Two transgenic lines, L- and H-, were generated that carried cDNAs encoding the L- and H-chains of a mouse IgG mAb, respectively, under the control of the enhancer-linked sericin-1 promoter. | ễFEBS Journal Production of a recombinant mouse monoclonal antibody in transgenic silkworm cocoons Masashi lizuka1 Shingo Ogawa2 Atsushi Takeuchi1 Shinichi Nakakita3 Yuhki Kubo4 Yoshitaka Miyawaki4 Jun Hirabayashi3 and Masahiro Tomita1 1 Neosilk Co. Ltd Higashihiroshima Hiroshima Japan 2 Research Institute Koken Co. Ltd Kita-ku Tokyo Japan 3 Life Science Research Center Kagawa University Kita-gun Kagawa Japan 4 Masuda ChemicalIndustries Co. Ltd Takamatsu Japan Keywords IgG monoclonalantibody N-glycosylation recombinant protein silk gland Correspondence M. Tomita Neosilk Co. Ltd 3-13-26 Kagamiyama Higashihiroshima Hiroshima 739-0046 Japan Fax 81 82 431 0654 Tel 81 82 431 0652 E-mail mtomita@ Received 15 June 2009 revised 3 August 2009 accepted 5 August 2009 doi In the present study we describe the production of transgenic silkworms expressing a recombinant mouse mAb in their cocoons. Two transgenic lines L- and H- were generated that carried cDNAs encoding the L- and H-chains of a mouse IgG mAb respectively under the control of the enhancer-linked sericin-1 promoter. Cocoon protein analysis indicated that the IgG L- or H-chain was secreted into the cocoons of each line. We also produced a transgenic line designated L H which carried both cDNAs by crossing the L- and H-lines. This line efficiently produced the recombinant mAb as a fully assembled H2L2 tetramer in its cocoons with negligible L- or H-chain monomer and H-chain dimer production. Thus the H2L2 tetramer was synthesized in and secreted from the middle silk gland cells. Crossing of the L H-line with a transgenic line expressing a baculovirus-derived trans-activator produced a increase in mAb expression. The recombinant mAb was extracted from the cocoons with a buffer containing 3 M urea and purified by protein G affinity column chromatography. The antigen-binding affinity of the purified recombinant mAb was identical to that of the native mAb produced by

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