Facile methods for generating human somatic cell gene knockouts using recombinant adeno-associated viruses
Facile methods for generating human somatic cell gene knockouts using recombinant adeno-associated viruses
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DOI：
10.1093/nar/gnh009
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发表时间：
2004-01-01
影响因子：
14.9
通讯作者：
Vogelstein, B
中科院分区：
文献类型：
--
作者：
Kohli, M;Rago, C;Vogelstein, B
新出现的证据表明，重组腺相关病毒（rAAV）载体可用于在人体细胞中的特异性基因靶向。我们已经开发了一种rAAV载体构建程序，采用融合PCR和一个克隆步骤，大大简化了敲除过程。我们通过破坏人类结肠癌细胞内特定位置的基因以及永生化正常上皮细胞内的基因来证明其效用。这项技术应广泛适用于需要操纵人类基因组的体外研究。
Emerging evidence suggests that recombinant adeno-associated viral (rAAV) vectors can be used for specific gene targeting in human somatic cells. We have developed an rAAV vector construction procedure employing fusion PCR and a single cloning step that considerably simplifies the knockout process. We demonstrate its utility by disrupting genes at specific positions within human colon cancer cells as well as within immortalized normal epithelial cells. This technology should be broadly applicable to in vitro studies that require the manipulation of the human genome.