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    Optimization of cells for endogenous gene activation
    8.
    发明授权
    Optimization of cells for endogenous gene activation 有权
    内源基因激活的细胞优化

    公开(公告)号：US07008764B1

    公开(公告)日：2006-03-07

    申请号：US09203500

    申请日：1998-12-01

    申请人： Konrad Honold ,  Thomas Holtschke ,  Anne Stern

    发明人： Konrad Honold ,  Thomas Holtschke ,  Anne Stern

    IPC分类号： C12Q1/68

    CPC分类号： C07K14/505 ,  C12N15/63 ,  C12N15/65 ,  C12N15/85 ,  C12N15/90 ,  C12N15/907 ,  C12N2800/30 ,  C12N2830/002 ,  C12Q1/6897

    摘要： The invention concerns a process for optimizing the gene expression in cells. A first aspect concerns a process for changing the expression of a nucleic acid sequence which is present endogenously in a eukaryotic cell by introduction of a heterologous expression control sequence into the genome of the cell by means of homologous recombination as well as site-specific recombinase-mediated excision of inserted foreign DNA and its replacement by further heterologous expression control sequences or/and amplification genes. In addition the invention concerns the introduction of one or several nucleic acid sequences to which an activator protein or an activator protein complex binds e.g. a hypoxia-inducible factor (HIF), into the genome of a eukaryotic cell by homologous recombination in order to change the expression of a target gene. Furthermore the invention concerns a process for testing the influence of 5′ or 3′ non-coding nucleic acid fragments on the expression of a target gene by determining the expression of a reporter gene. In addition the invention concerns a process for providing a DHFR-negative eukaryotic cell containing a recombinase target sequence as well as the expression of a nucleic acid sequence inserted into the recombinase target sequence.

    摘要翻译： 本发明涉及优化细胞中基因表达的方法。 第一方面涉及一种通过将异源表达控制序列通过同源重组以及位点特异性重组酶导入到细胞基因组中而在真核细胞中内源性存在的核酸序列的表达的方法。 介导的插入的外源DNA的切除及其通过进一步的异源表达控制序列或/和扩增基因的替代。 此外，本发明涉及一种或几种核酸序列的引入，活化蛋白或活化蛋白复合物例如与其结合。 缺氧诱导因子（HIF），通过同源重组进入真核细胞的基因组，以改变靶基因的表达。 此外，本发明涉及通过测定报告基因的表达来测试5'或3'非编码核酸片段对靶基因表达的影响的方法。 此外，本发明涉及提供含有重组酶靶序列的DHFR阴性真核细胞以及插入重组酶靶序列中的核酸序列的表达的方法。