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    COMPOSITION AND METHOD FOR DIVERSIFICATION OF TARGET SEQUENCES
    8.
    发明公开
    COMPOSITION AND METHOD FOR DIVERSIFICATION OF TARGET SEQUENCES 审中-公开
    目标序列多样性的组成和方法

    公开(公告)号：EP2825652A1

    公开(公告)日：2015-01-21

    申请号：EP13761578.7

    申请日：2013-03-13

    申请人： Omeros Corporation ,  University Of Washington Through Its Center For Commercialization

    发明人： CUMMINGS, W., Jason ,  TJOELKER, Larry, W. ,  WOOD, Christi, L. ,  YABUKI, Munehisa ,  ALLISON, Daniel, S. ,  LEPPARD, John, B. ,  MAIZELS, Nancy

    IPC分类号： C12N15/63 ,  C12N15/13 ,  C12N15/70 ,  C12N15/79

    CPC分类号： C12N15/1082 ,  C07K16/00 ,  C07K16/2863 ,  C07K16/30 ,  C07K2317/24 ,  C07K2317/92

    摘要： The disclosure relates generally to the targeting of genes to, and their integration into, an innnnunoglobulin (antibody) heavy chain locus. In particular, the methods described herein contemplate replacing the single rearranged heavy chain V, D, and J genes of a B cell lymphoma such as DT40 with independently rearranged V
    H -D-J
    H genes of chicken, in a system for generating immunoglobulin diversity. Also contemplated is replacement of the chicken V
    H -D-J
    H with rearranged V
    H -D-J
    H genes of other vertebrates including human in a system for generating immunoglobulin diversity, with the exception of any substitution disclosed and claimed in PCT application WO 2009/029315 A2. Regions upstream of the rearranged V
    H gene, and downstream of the rearranged J
    H genes, are identified and incorporated into the immunoglobulin heavy chain gene-targeting vectors for efficient gene replacement. Also described is construction of a diverse chicken immunoglobulin heavy chain VDJ library in DT40 by homologous gene replacement of the single endogenous rearranged VDJ gene with a chicken VDJ repertoire using the described targeting vectors. The methods are useful for generating a diverse library of immunoglobulins that can be screened for binding to desired target antigens. Compositions include the unique targeting vectors, the diverse chicken heavy chain VDJ library in the targeting vectors, and the diverse chicken VDJ DT40 library that is created by homologous gene replacement of the single endogenous rearranged VDJ gene with a chicken VDJ repertoire, using the targeting vectors.

    摘要翻译： 本公开内容一般涉及基因靶向以及它们整合入免疫球蛋白（抗体）重链基因座。 具体而言，本文所述的方法考虑在用于产生免疫球蛋白多样性的系统中用鸡的独立重排的V H-D-J H基因置换B细胞淋巴瘤例如DT40的单重排的重链V，D和J基因。 除了在PCT申请WO 2009/029315 A2中公开并要求保护的任何替代之外，还考虑用在用于产生免疫球蛋白多样性的系统中的其他脊椎动物包括人的重排V H -D-J H基因置换鸡V H-D-J H。 重排的V H基因的上游区域和重排的J H基因的下游区域被鉴定并掺入到免疫球蛋白重链基因靶向载体中用于有效的基因置换。 还描述了在DT40中构建多样的鸡免疫球蛋白重链VDJ文库，其通过使用所述靶向载体用鸡VDJ库同源基因置换单一内源重排VDJ基因。 该方法可用于产生可筛选与所需靶抗原结合的免疫球蛋白的多样文库。 组合物包括独特的靶向载体，靶向载体中的不同鸡重链VDJ文库和通过用鸡VDJ库同源基因替换单一内源重排VDJ基因产生的不同鸡VDJ DT40文库，使用靶向载体 。