Targeted recombination at the Chinese hamster APRT locus using insertion versus replacement vectors.
Targeted recombination at the Chinese hamster APRT locus using insertion versus replacement vectors.
复制标题
使用插入与替换载体在中国仓鼠 APRT 基因座上进行靶向重组。
DOI：
10.1023/b:scam.0000007112.62928.d8
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Nairn,RS
中科院分区：
文献类型：
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作者：
Adair,GM;Scheerer,JB;Brotherman,A;McConville,S;Wilson,JH;Nairn,RS
在本研究中，我们研究了靶向载体构型和载体线性化位点对内源CHOAPRT基因座靶向重组频率的影响，并分析了在APRT靶向实验中获得的APRT+重组体的类型和类别分布，所述靶向实验使用未切割的环状、插入型（末端插入）和重叠型（末端插入）。相同pAG 7靶向载体的末端向外（ends-out）构型，包括通过在APRT靶向同源性内或5′或3′边界处的位点处引入双链断裂（DSB）产生的构型。我们的研究结果表明：1）哺乳动物细胞中的质粒-染色体靶向重组可能不会被靶向载体中的DSB刺激到与染色体靶标中的DSB相同的程度; 2）重组类别分布高度依赖于靶向载体构型;和3）单侧侵入机制可能在哺乳动物细胞中的同源重组中起重要作用。
In this study, we have examined the effects of targeting vector configuration and site of vector linearization on the frequency of targeted recombination at the endogenous CHOAPRTlocus, and have analyzed the types and class distributions of APRT+recombinants obtained inAPRTtargeting experiments employing uncut circular, insertion-type (ends-in), and replacement-type (ends-out) configurations of the same pAG7 targeting vector, including configurations produced by introduction of a double-strand break (DSB) at sites either within, or at the 5′ or 3′ boundaries ofAPRTtargeting homology. Our results suggest that: 1) plasmid-chromosome targeted recombination in mammalian cells may not be stimulated to the same degree by a DSB in the targeting vector as by a DSB in the chromosomal target; 2) recombinant class distributions are highly dependent upon targeting vector configuration; and 3) one-sided invasion mechanisms may play a significant role in homologous recombination in mammalian cells.