A Simple and Convenient Sticky/Blunt-End Ligation Method for Fusion Gene Construction
A Simple and Convenient Sticky/Blunt-End Ligation Method for Fusion Gene Construction
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一种简单方便的融合基因构建粘/平端连接方法
DOI：
10.1007/s10528-015-9669-x
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发表时间：
2015-04-01
影响因子：
2.4
通讯作者：
Zhang, Qingrui
中科院分区：
文献类型：
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作者：
Gao, Song;Zhang, Jiannan;Zhang, Qingrui
在这里，我们提出了一种简单便捷的粘性/钝连接方法，用于融合基因构建。融合基因是通过无缝连接5'端磷酸化钝器末端而不是由重叠延伸PCR（OE-PCR）构建的。因此，可以避免通过PCR融合通过两个单功能酶基因构建的大型DNA片段（例如，大型双功能酶基因）的挑战。此外，不需要对OE-PCR的内引物的合成，这表明该方法对于构建具有多个片段组合的融合基因特别方便（例如，嵌合基因库，融合基因库）。作为对常用融合基因构建技术的修改，该方法可能在生物科学和生物技术中发现了广泛的应用。
Here we present a simple and convenient sticky/blunt-end ligation method for fusion gene construction. The fusion gene is constructed by seamless ligation of 5'-end phosphorylated blunt ends instead of by overlap extension PCR (OE-PCR). Therefore, the challenge of amplifying large DNA fragments (e.g., the large bifunctional enzyme gene constructed by fusion of two monofunctional enzyme genes) by PCR can be avoided. In addition, synthesis of the inner primers for OE-PCR is not necessary, indicating that this method should be especially convenient for construction of fusion genes with various combinations of multiple fragments (e.g., chimeric gene libraries, fusion gene libraries). As a modification of the commonly used fusion gene construction technique, this method may find a wide range of applications in bioscience and biotechnology.