利用插入载体和替代载体对中国仓鼠APRT位点进行靶向重组。

G M Adair, J B Scheerer, A Brotherman, S McConville, J H Wilson, R S Nairn
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引用次数: 16

摘要

在本研究中,我们研究了靶向载体配置和载体线性化位点对内源性CHO APRT位点靶向重组频率的影响,并分析了采用同一pAG7靶向载体的未切割圆形、插入型(端入)和替换型(端出)配置的APRT+重组体在APRT靶向实验中获得的类型和类别分布。包括通过在APRT靶向同源性的5'或3'边界内的位点引入双链断裂(DSB)而产生的构型。结果表明:1)靶载体中的DSB对哺乳动物细胞质粒染色体靶向重组的刺激程度可能与靶染色体中的DSB不同;2)重组类分布高度依赖于靶向载体配置;3)单侧侵袭机制可能在哺乳动物细胞同源重组中发挥重要作用。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Targeted recombination at the Chinese hamster APRT locus using insertion versus replacement vectors.

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 CHO APRT locus, and have analyzed the types and class distributions of APRT+ recombinants obtained in APRT targeting 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 of APRT targeting 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.

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