DEVELOPMENT OF AGROBACTERIUM-BASED TRANSIENT GENE EXPRESSION ASSAY IN ROSE LEAVES

Abstract

Post-genomics need tools to analyze genomic data for its functional characterization within short time. Agrobacterium mediated transient gene expression assays are popular choice to analyze the function of a gene within a few days. These transient gene expression assays have already proved their utility in different plant species Here we explored the affect of different physical, chemical and biological parameters on transient gene expression in rose leaves using β-glucuronidase (GUS) reporter gene. The selected rose genotypes for this study were Pariser Charme, 91/100-5 and 88/124-46 which were maintained in three different pre-culturing conditions i.e. climate chamber, in vitro culture and/ or in tunnels. The leaves of these cultivars were infiltrated with different densities of Agrobacterium strain GV3101 and the assay was optimized. The efficiency of this assay was found to mainly depend on rose genotypes, age of leaves, plant pre-culture method and density of Agrobacterium. The highest GUS expression was recorded in type B leaves (light green leaves with dark green prominent veins and red edges) of Pariser Charme and type C leaves (complete light green young leaves) of 91/100-5 grown in tunnels and when kept at 22o C in dark after agro-infiltration. The optimized bacterial density for the best GUS expression was found to be OD600 - 1.5 in simple water without acetosyringone.