Nonreciprocal exchanges of information between DNA duplexes coinjected into mammalian cell nuclei
Nonreciprocal exchanges of information between DNA duplexes coinjected into mammalian cell nuclei
复制标题
共注射到哺乳动物细胞核中的 DNA 双链体之间的非互易信息交换
DOI：
10.1128/mcb.5.1.59-69.1985
复制
发表时间：
1985
影响因子：
5.3
通讯作者：
Mario R. Capecchi
中科院分区：
文献类型：
--
作者：
K. Folger;K. Thomas;Mario R. Capecchi
我们研究了共注射到培养的哺乳动物细胞中的质粒分子之间的同源重组机制。通过从Neor基因中携带不同琥珀突变的两种质粒中选择重建功能性Neor基因，获得含有重组DNA分子的细胞系。此外，这些质粒在Neor基因内和附近含有限制性长度多态性。这些多态性不赋予可选择的表型，但用于鉴定和分类选择和非选择的重组DNA分子。从该分析得出的惊人结论是，在短距离内共注射的质粒分子之间交换信息的主要机制（即，少于1个内切酶）通过非相互同源重组进行。在培养的哺乳动物细胞中，共注射的质粒分子之间的同源重组频率非常高，接近统一。我们证明，这种高频率既不需要每个细胞的质粒分子的高输入，也不需要细胞核内质粒DNA的局部高浓度。因此，似乎质粒分子一旦被引入细胞核，即使在存在大量过量的宿主DNA序列的情况下，也可以毫不费力地相互寻找并参与同源重组。最后，我们表明，大多数的同源重组事件发生在1小时的间隔后，质粒DNA引入细胞核。
We have examined the mechanism of homologous recombination between plasmid molecules coinjected into cultured mammalian cells. Cell lines containing recombinant DNA molecules were obtained by selecting for the reconstruction of a functional Neor gene from two plasmids that bear different amber mutations in the Neor gene. In addition, these plasmids contain restriction-length polymorphisms within and near the Neor gene. These polymorphisms did not confer a selectable phenotype but were used to identify and categorize selected and nonselected recombinant DNA molecules. The striking conclusion from this analysis is that the predominant mechanism for the exchange of information between coinjected plasmid molecules over short distances (i.e., less than 1 kilobase) proceeds via nonreciprocal homologous recombination. The frequency of homologous recombination between coinjected plasmid molecules in cultured mammalian cells is extremely high, approaching unity. We demonstrate that this high frequency requires neither a high input of plasmid molecules per cell nor a localized high concentration of plasmid DNA within the nucleus. Thus, it appears that plasmid molecules, once introduced into the nucleus, have no difficulty seeking each other out and participating in homologous recombination even in the presence of a vast excess of host DNA sequences. Finally, we show that most of the homologous recombination events occur within a 1-h interval after the introduction of plasmid DNA into the cell nucleus.
使用腺病毒重叠末端DNA片段进行重组的遗传分析。
影响因子：
3.7
作者：
Volkert,FC;Young,CS
通讯作者：
Young,CS
哺乳动物细胞中的同源重组介导两个重叠基因片段形成功能基因。
DOI：
10.1073/pnas.80.7.2002
发表时间：
1983
影响因子：
11.1
作者：
deSaintVincent,BR;Wahl,GM
通讯作者：
Wahl,GM