摘要
随着越来越多基因组全序列的测定完成,基因组的研究进入了功能研究阶段。功能基因组的研究需要把大量基因连入不同载体,传统的酶切连接构建载体的方法不能满足这种大规模克隆的需要。Gateway技术是一种高效的大规模克隆系统,而且对载体和宿主没有依赖性。本文利用Gateway大规模克隆技术将16个拟南芥转录因子的ORF克隆入植物表达载pPTV和酵母表达载体pYTV中,酵母融合表达实验和West-ern-blot检测证明了该克隆途径的可行性,并且得到的His-Tag融合蛋白,为拟南芥转录因子的大规模蛋白质组学研究奠定了良好的基础,同时基因序列的聚类分析为将来进一步的功能研究提供了有利信息。
With the completion of more and more genome sequences, genomic research has gone into the functional genomic era. To identify genome functions, a great many genes need to be cloned into different vectors, while conventional method with restriction digestion and ligation appeared powerless in large scale cloning. Gateway cloning technology is a fast and reliable large scale cloning system which is independent of the host cells and vectors. In this article, 16 transcription factors of Arabidopsis were cloned into plant expression vector pPTV and yeast expression vector pYTV using Gateway cloning technology. Most of them were expressed correctly in yeast by western blot detection, which shows Gateway cloning technology is effective in high-throughput cloning. The His-tag fusion proteins of transcription factors we got provided the possibility for large-scale proteinomic research and the clustering analysis of protein sequences was useful for further functional research.
出处
《分子植物育种》
CAS
CSCD
2004年第3期358-364,共7页
Molecular Plant Breeding
基金
国家自然基金委员会重大国际合作项目(30221120261)资助。
