Targeted engineering of the Caenorhabditis elegans genome following Mos1-triggered chromosomal breaks
Targeted engineering of the Caenorhabditis elegans genome following Mos1-triggered chromosomal breaks
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DOI：
10.1038/sj.emboj.7601463
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发表时间：
2007-01-10
期刊：
影响因子：
11.4
通讯作者：
Bessereau, Jean-Louis
中科院分区：
文献类型：
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作者：
Robert, Valerie;Bessereau, Jean-Louis
果蝇元件 Mos1 是一种 II 类转座子，通过“剪切和粘贴”机制移动，并且可以通过实验在秀丽隐杆线虫种系中进行动员。在这里，我们触发了已识别的 Mos1 插入的切除，以在给定位点产生染色体断裂并进一步操纵断裂的位点。双链断裂（DSB）修复可以通过使用含有与断裂染色体区域同源的序列的转基因作为修复模板的基因转换来实现。因此，转基因中设计的突变可以高频率复制到特定基因座。对该途径进行了进一步的表征，以开发一种有效的工具（称为 MosTIC）来操纵线虫基因组。 MosTIC 实验期间对 DSB 修复的分析表明，DSB 也可以通过种系中的末端连接来密封，独立于进化上保守的 Ku80 和连接酶 IV 因子。结合目前正在生成的公开可用的 Mos1 插入库，MosTIC 将提供定制线虫基因组的通用工具。
The Drosophila element Mos1 is a class II transposon, which moves by a 'cut-and-paste' mechanism and can be experimentally mobilized in the Caenorhabditis elegans germ line. Here, we triggered the excision of identified Mos1 insertions to create chromosomal breaks at given sites and further manipulate the broken loci. Double-strand break (DSB) repair could be achieved by gene conversion using a transgene containing sequences homologous to the broken chromosomal region as a repair template. Consequently, mutations engineered in the transgene could be copied to a specific locus at high frequency. This pathway was further characterized to develop an efficient tool - called MosTIC - to manipulate the C. elegans genome. Analysis of DSB repair during MosTIC experiments demonstrated that DSBs could also be sealed by end-joining in the germ line, independently from the evolutionarily conserved Ku80 and ligase IV factors. In conjunction with a publicly available Mos1 insertion library currently being generated, MosTIC will provide a general tool to customize the C. elegans genome.