用于快速分离增强子的条件性复制分析（CRA）程序。
Contingent replication assay (CRA) procedure for rapid isolation of enhancers.
作者信息
Vasavada H A, Lengyel P, Weissman S M
出版信息
Gene. 1987;55(1):29-40. doi: 10.1016/0378-1119(87)90245-9.
A rapid procedure for the isolation of functional enhancer sequences consists of the construction of a shotgun DNA library in SV40-based plasmid shuttle vectors which depend on an enhancer for replication, the replication in monkey (CVI) cells of those vectors into which an enhancer sequence was inserted, the selective cleavage of unreplicated vectors by DpnI and the recovery of the replicated vectors by transfection into Escherichia coli. We describe conditions for the fusion of protoplasts to CVI cells, under which conditions the probability of only one type of plasmid entering a cell is increased and thus complementation and rescue of enhancer-less plasmids are decreased. The effectiveness of the procedure is demonstrated by the recovery of enhancers from bovine papillomavirus and Moloney murine sarcoma virus.
一种分离功能性增强子序列的快速方法包括
在基于SV40的质粒穿梭载体中构建鸟枪法DNA文库，该载体依赖增强子进行复制；将插入了增强子序列的载体在猴（CVI）细胞中进行复制；用DpnI选择性切割未复制的载体；通过转染大肠杆菌回收复制的载体。我们描述了原生质体与CVI细胞融合的条件，在此条件下，只有一种质粒进入细胞的概率增加，从而减少了无增强子质粒的互补和拯救。从牛乳头瘤病毒和莫洛尼鼠肉瘤病毒中回收增强子证明了该方法的有效性。
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