Plant Diversity ›› 2012, Vol. 34 ›› Issue (4): 397-.doi: 10.3724/SP.J.1143.2012.11185

• Articles • Previous Articles     Next Articles

Simplification of Entry Vector by TA Approach

 YAN  Xian-Lun-1、2, WANG  Chun-Tao-1 , KONG  Xiang-Xiang-1, YANG  Yong-Ping-1, HU  Xiang-Yang-1   

  1. 1 Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650204, China;
    2 Graduate University of Chinese Academy of Sciences, Beijing 100049, China
  • Received:2011-12-14 Online:2012-08-25 Published:2012-04-01
  • Supported by:

    The Major Science and Technology Program (110201101003TS03, 2011YN02和2011YN03)

Abstract:

Gateway technology is a universal cloning approach that enables rapid cloning of DNA fragments into multiple Gatewaycompatible destination vectors using λ phage sitespecific recombination, eliminating the requirement to work with restriction enzymes and ligase. But a problem using this system for making entry clone is the expensiveness and long time to buy the enzyme. To solve this problem, we created the TA cloning entry vector that contained a Ttail in each 3’end through modification of pDONR207. The TA cloning approach can construct entry clones simply, economically and rapidly. Using Gateway T vectors prepared by this improved method, prokaryotic expression vector and eukaryotic expression vector for SOS2 gene were constructed. Through the methods of prokaryotic expression and transient gene expression in Arabidopsis protoplasts, it proved that the SOS2 gene expressed well in both prokaryotic cells and eukaryotic cells.

Key words: Gateway technology, TA cloning, Entry clone, Prokaryotic expression, Transient gene expression in Arabidopsis protoplasts

CLC Number: 

  • Q 943