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RMDAP: A Versatile, Ready-To-Use Toolbox for Multigene Genetic Transformation

Lei Ma,Jiangli Dong,Yongsheng Jin,Mingliang Chen,Xiaoye Shen,Tao Wang

Published 2011 in PLoS ONE

ABSTRACT

Background The use of transgenes to improve complex traits in crops has challenged current genetic transformation technology for multigene transfer. Therefore, a multigene transformation strategy for use in plant molecular biology and plant genetic breeding is thus needed. Methodology/Principal Findings Here we describe a versatile, ready-to-use multigene genetic transformation method, named the Recombination-assisted Multifunctional DNA Assembly Platform (RMDAP), which combines many of the useful features of existing plant transformation systems. This platform incorporates three widely-used recombination systems, namely, Gateway technology, in vivo Cre/loxP and recombineering into a highly efficient and reliable approach for gene assembly. RMDAP proposes a strategy for gene stacking and contains a wide range of flexible, modular vectors offering a series of functionally validated genetic elements to manipulate transgene overexpression or gene silencing involved in a metabolic pathway. In particular, the ability to construct a multigene marker-free vector is another attractive feature. The built-in flexibility of original vectors has greatly increased the expansibility and applicability of the system. A proof-of-principle experiment was confirmed by successfully transferring several heterologous genes into the plant genome. Conclusions/Significance This platform is a ready-to-use toolbox for full exploitation of the potential for coordinate regulation of metabolic pathways and molecular breeding, and will eventually achieve the aim of what we call “one-stop breeding.”

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