The application of the Cre-loxP system for generating multiple knock-out and knock-in targeted loci.
The application of the Cre-loxP system for generating multiple knock-out and knock-in targeted loci.
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DOI：
10.1007/978-1-62703-302-2_13
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发表时间：
2013-01-01
期刊：
影响因子：
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通讯作者：
Kimmel, Alan R
中科院分区：
文献类型：
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作者：
Faix, Jan;Linkner, Joern;Kimmel, Alan R
盘基网柄藻是研究生长、发育和基本细胞过程的一种非常有效的真核生物模型。其小尺寸的单倍体基因组允许高效的靶向同源重组，用于基因破坏和敲入表位标记。我们先前描述了一个强大的系统，用于在网柄藻中产生多个基因突变，通过在Cre重组酶瞬时表达后再循环杀稻瘟菌素S选择标记。我们现在已经进一步优化了该系统以获得更高的效率，此外，还将其与敲除和敲入基因靶向相结合，从而可以在单个细胞系中表征多个和协作基因功能。
Dictyostelium discoideum is an exceptionally powerful eukaryotic model to study many aspects of growth, development, and fundamental cellular processes. Its small-sized, haploid genome allows highly efficient targeted homologous recombination for gene disruption and knock-in epitope tagging. We previously described a robust system for the generation of multiple gene mutations in Dictyostelium by recycling the Blasticidin S selectable marker after transient expression of the Cre recombinase. We have now further optimized the system for higher efficiency and, additionally, coupled it to both, knock-out and knock-in gene targeting, allowing the characterization of multiple and cooperative gene functions in a single cell line.