Targeted mutagenesis by homologous recombination in D-melanogaster
Targeted mutagenesis by homologous recombination in D-melanogaster
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DOI：
10.1101/gad.986602
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发表时间：
2002-06-15
影响因子：
10.5
通讯作者：
Golic, KG
中科院分区：
文献类型：
--
作者：
Rong, YKS;Titen, SW;Golic, KG
我们使用最近开发的方法来产生果蝇五个内源基因的突变等位基因，包括p53肿瘤抑制基因的同源物。FLP位点特异性重组酶和I-SceI核酸内切酶的转基因表达在体内产生染色体外线性DNA分子。这些分子与相应的染色体基因座进行同源重组，以产生宿主基因组的靶向改变。结果解决了几个问题，这种技术的一般效用。我们表明，端粒附近的基因可以有效地靶向，没有知识的突变表型是需要靶向，插入突变和等位基因取代可以很容易地产生。
We used a recently developed method to produce mutant alleles of five endogenous Drosophila genes, including the homolog of the p53 tumor suppressor. Transgenic expression of the FLP site-specific recombinase and the I-SceI endonuclease generates extrachromosomal linear DNA molecules in vivo. These molecules undergo homologous recombination with the corresponding chromosomal locus to generate targeted alterations of the host genome. The results address several questions about the general utility of this technique. We show that genes not near telomeres can be efficiently targeted; that no knowledge of the mutant phenotype is needed for targeting; and that insertional mutations and allelic substitutions can be easily produced.