A Simple PCR Cloning Step to Utilize a Binary Vector for Identification and Characterization of Enhancer Element in Plant Promoters

Abstract

Identification and characterization of regulatory elements, such as enhancers, require designing of specialized plant transformation vector to testify its unique features of position and orientation-independent core-promoter activation. Very few plant transformation vectors are available till date for identification of enhancer elements in plant promoters. Hence, we need to utilize the existing binary plant transformation vectors, such as pBI101, through tedious and time-consuming restriction enzyme-based cloning steps. We describe here a simple one-step PCR-based cloning strategy to introduce a 35S CaMV minimal promoter in a binary plant transformation vector pBI101, and utilize the unique features to identify a lateral organ junction tissue-specific enhancer element in a plant promoter from Arabidopsis thaliana. © 2012 NAAS (National Academy of Agricultural Sciences).