The current status of plant transformation technologies

Abstract

Plant transformation vectors and methodologies have been improved to increase the efficiency of plant trans - formation and to achieve stable expression of transgenes in plan ts. Due to the simplicity of the transformation system and precise integration of transgenes, Agro- bacterium Ti plasmid -based vectors continue to offer the best system for plant transformation. Binary vectors have been improved by the incorporation of supe rvirulent vir genes, matrix attachment regions (MAR) and the inser- tion of introns in marker genes and reporter genes. With these improvements and with the use of acetosyringon e, transformation of monocotyledon ous plants using Agro- bacterium has almost become a routine process. The green fluorescent protein (GFP) gene has been extensively modified for plant codon preference, ER/plastid targeting and for greater solubility thereby making it a versatile vital reporter for transgenic plants. Significant progr ess is seen in developing transgenic plants devoid of antibiotic marker genes. Cotransformation of multiple T-DNAs, site-specific recombination strategies and deploy ment of Ac/Ds-based transposition have helped in the elimination of marker genes in transg enic plants. Positive selection strategies using ipt, xylose isomerase and phospho-mannose isomerase have been demonstrated to be useful in many crop plants. The development of BIBAC vectors, a demonstrated capability to transfer multiple genes of a pathway and successful T-DNA tagging in rice, signal the readiness with which transformation technologies can be deployed for the study of 'functional genomics' in plants. The particle bombardment system continues to find use in organelle transformation and tran sformation of plants that lack efficient regeneration systems. A detailed understanding of gene silencing has led to the design of vectors that minimize transgene silencing while ensuring desired levels of transgene expression. Efforts are underway to unde rstand the mechanism of T -DNA integration in plants so that 'knock out' mutagenesis and homology-based gene replacements can be achieved in plants. We review in this article the current status of transformation technologies. An overview of the status of deployment of plant transformation technologies in India is also presented. Plant transformation is performed using a wide range of tools such as Agrobacterium Ti plasmid vectors, microprojectile bombardment, microinjection, chemical (PEG) treatment of prot oplasts and electroporation of protoplasts. Though all methods have advantages that are unique to each of them, transformation using Agrobac- terium and microprojectile bombardment are currently the most extensively used methods 1 . Recent develop- ments in t hese two technologies have been reviewed together with the phenomenon of 'gene silencing' that has come to centrestage after a large number of trans - genic plants have been carefully evaluated for transgene expression in successive generations.