Using Multisite LR Cloning to Generate a Destination Clone.
Using Multisite LR Cloning to Generate a Destination Clone.
复制标题
使用多站点 LR 克隆生成目标克隆。
DOI：
10.1101/pdb.prot094946
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发表时间：
2018
影响因子：
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通讯作者：
Walhout,AlberthaJM
中科院分区：
文献类型：
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作者：
Reece-Hoyes,JohnS;Walhout,AlberthaJM
This protocol describes using the Gateway recombinatorial cloning system to simultaneously transfer a promoter and an open reading frame (ORF) from two different Entry clones into the same Destination vector using LR enzymes. A multisite cloning reaction transfers the inserts from multiple Entry clones into a single Destination vector. This type of recombination is much less efficient than transferring a single DNA fragment; however, the variety of Destination clones that can be generated in this manner is vast. In this example protocol, we describe using pDEST-MB14 to make a Destination clone that features a promoter fragment fused upstream to an ORF that is cloned in-frame with a carboxy-terminal green fluorescent protein (GFP) moiety encoded by the plasmid backbone. This method can be used as a guide for other multisite cloning reactions.
