USER fusion: a rapid and efficient method for simultaneous fusion and cloning of multiple PCR products.
USER fusion: a rapid and efficient method for simultaneous fusion and cloning of multiple PCR products.
复制标题
用户融合：一种快速有效的方法，用于同时融合和克隆多个PCR产品。
DOI：
10.1093/nar/gkm106
复制
发表时间：
2007
影响因子：
14.9
通讯作者：
Halkier, Barbara A.
中科院分区：
文献类型：
--
作者：
Geu-Flores, Fernando;Nour-Eldin, Hussam H.;Nielsen, Morten T.;Halkier, Barbara A.
我们提出了一种方法，允许同时融合和克隆的多个PCR产物在一个快速和有效的方式。该方法基于使用在其5′端附近含有单个脱氧尿苷残基的PCR引物。用商业脱氧尿苷切除试剂处理PCR产物产生长的3′突出端，其设计为特异性互补。将该原理与改进的USER克隆技术相结合，提供了一种简单、快速且非常有效的方法来同时将多个PCR片段融合并克隆到感兴趣的载体中。当三种不同的PCR产物融合并克隆到用户兼容的载体中时，在一个简单的程序中获得了约90%的阳性克隆，除了单个PCR扩增步骤和细菌转化外，大约需要一个小时。我们希望这种方法可以取代重叠PCR和IIS型限制性内切酶在许多应用中的使用。
We present a method that allows simultaneous fusion and cloning of multiple PCR products in a rapid and efficient manner. The procedure is based on the use of PCR primers that contain a single deoxyuridine residue near their 5′ end. Treatment of the PCR products with a commercial deoxyuridine-excision reagent generates long 3′ overhangs designed to specifically complement each other. The combination of this principle with the improved USER cloning technique provides a simple, fast and very efficient method to simultaneously fuse and clone multiple PCR fragments into a vector of interest. Around 90% positive clones were obtained when three different PCR products were fused and cloned into a USER-compatible vector in a simple procedure that, apart from the single PCR amplification step and the bacterial transformation, took approximately one hour. We expect this method to replace overlapping PCR and the use of type IIS restriction enzymes in many of their applications.
登录
查看更多内容
影响因子：
4.8
作者：
Jiang, DY;Hatahet, Z;Wallace, SS
通讯作者：
Wallace, SS
影响因子：
7.7
作者：
Rashtchian, Ayoub
通讯作者：
Rashtchian, Ayoub
影响因子：
2.7
作者：
Watson, DE;Bennett, GN
通讯作者：
Bennett, GN
影响因子：
14.9
作者：
Nour-Eldin, Hussam H.;Hansen, Bjarne G.;Norholm, Morten H. H.;Jensen, Jacob K.;Halkier, Barbara A.
通讯作者：
Halkier, Barbara A.
影响因子：
3.5
作者：
HORTON, RM;HUNT, HD;PEASE, LR
通讯作者：
PEASE, LR