EXPRESSION ANALYSIS OF ANTI-SENSE O S PPO GENE PROMOTER IN RESPONSE TO PLANT GROWTH HORMONES IN TRANSGENIC MODEL PLANT

Abstract

Polyphenol oxidases (PPOs) are ubiquitous copper containing enzymes, involved in the oxidation of ortho-diphenolics into quinones. They are broadly distributed and extensively studied oxidative enzymes, playing pivotal role against various biotic and abiotic stresses. Purpose of the present study was to examine the activity of antisense OsPPO promoter in response to exogenous applications of α-naphthaline acetic acid (NAA), 6-benzyl amino purine (BAP) and abscissic acid (ABA). Tobacco plants were transformed with antisense OsPPO promoter, fused with GUS reporter gene using Agrobacterium mediated transformation and transformation was confirmed by PCR and histochemical analysis. GUS expression pattern of antisense OsPPO promoter was analysed in response to various concentrations of NAA (0.1 μM, 1 μM, 10 μM and 100 μM), BAP (1 μM, 20 μM, 40 μM and 80 μM) and ABA (50 μM, 150 μM, 250 μM and 350 μM). It was found that antisense OsPPO promoter was strongly induced in response to different concentrations of these hormones, however induction of promoter was increased with increasing concentration of hormone and timing of treatment upto certain level and then it was decreased. Among different concentrations of these hormones applied, maximum GUS expression was observed at 10 μM of NAA treatment after 12 hours. Moreover promoter was strongly induced in response to 80 μM of BAP application and 350 μM of ABA. Microscopic analysis showed GUS expression in various parts of leaves e.g. guard cells, midrib, and veins while expression was also observed in root hairs. The results revealed that antisense OsPPO promoter is responsive towards hormonal stresses and suggested the idea that antisense OsPPO gene promoter can be utilized in transformation experiments regarding biotic stresses carries considerable importance in crop biotechnology for generating resistant cultivar against various stresses.