Homology dependence of targeted recombination at the Chinese hamster APRT locus.
Homology dependence of targeted recombination at the Chinese hamster APRT locus.
复制标题
中国仓鼠 APRT 基因座靶向重组的同源依赖性。
DOI：
10.1128/mcb.14.10.6663-6673.1994
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发表时间：
1994
影响因子：
5.3
通讯作者：
Adair,GM
中科院分区：
文献类型：
--
作者：
Scheerer,JB;Adair,GM
以中国仓鼠腺嘌呤磷酸核糖转移酶（APRT）基因的简单线性片段为靶向载体，研究了内源性APRT基因在中国仓鼠卵巢（CHO）细胞中的同源重组依赖性。我们已经检查了改变靶向序列同源性的总长度或5′或3′侧翼同源性的长度对靶向同源重组的频率和获得的重组事件的类型的影响。我们发现，APRT靶向同源性的长度与CHOAPRT基因座处靶向重组的频率之间存在指数（对数）关系，靶向重组的频率取决于靶向同源性的总长度和靶基因缺失两侧的同源性长度。尽管分析的大多数APRT+重组体反映了靶基因缺失的简单靶向置换或转化，很大一部分似乎是由靶基因引起的，在靶基因缺失一侧侧翼具有有限靶向同源性的APRT片段产生成比例地更少的靶基因转换事件和成比例地更多的模板化延伸和载体校正事件而不是侧翼同源性更高的片段。
Using simple linear fragments of the Chinese hamster adenine phosphoribosyltransferase (APRT) gene as targeting vectors, we have investigated the homology dependence of targeted recombination at the endogenousAPRTlocus in Chinese hamster ovary (CHO) cells. We have examined the effects of varying either the overall length of targeting sequence homology or the length of 5′ or 3′ flanking homology on both the frequency of targeted homologous recombination and the types of recombination events that are obtained. We find an exponential (logarithmic) relationship between length ofAPRTtargeting homology and the frequency of targeted recombination at the CHOAPRTlocus, with the frequency of targeted recombination dependent upon both the overall length of targeting homology and the length of homology flanking each side of the target gene deletion. Although most of the APRT+recombinants analyzed reflect simple targeted replacement or conversion of the target gene deletion, a significant fraction appear to have arisen by target gene-templated extension and correction of the targeting fragment sequences.APRTfragments with limited targeting homology flanking one side of the target gene deletion yield proportionately fewer target gene conversion events and proportionately more templated extension and vector correction events than do fragments with more substantial flanking homology.