PHYSIOLOGICAL AND MOLECULAR STUDIES OF TOMATO UNDER SALINITY STRESS

Abstract

The aim of the present investigation was to determine the effect of salt stress on the physiology of twelve genotypes of tomato (Solanum lycopersicum L.), to study the genetic diversity among these genotypes and to transfer salt tolerant gene in the selected genotype through Agrobacterium mediated gene transformation method. The seeds of different tomato genotypes were obtained from Ayub Agricultural Research Institute (AARI), Faisalabad and from Horticulture Research institute (HRI), NARC, Islamabad. First experiment comprises study of plant growth, ionic content, ABA content of shoot and root and proline content at four different NaCl salinity level. The genotypes Avinash, NTH242 and Gol performed better at higher salt concentration as indicated by less inhibition of plant height, shoot and root fresh weight and shoot and root dry weight. There was less translocation of Na + to shoot but greater accumulation of K + and Ca 2+ concomitant with higher production of osmoregulant (proline) and endogenous ABA accumulation in shoot was recorded. Second experiment was conducted to study the genetic diversity among the twelve tomato genotypes through randomly amplified polymorphic DNA analyses. Twelve tomato genotypes were grouped in four clusters at similarity coefficient value of 0.88 on the basis of randomly amplified polymorphic DNA analysis. Cluster-I consisted of a single tomato genotype “NTH-242”. Cluster-II of three tomato genotypes Lyp No.1, BARI, Roma and Cluster-III grouped Pakit and Riogrande together. Cluster-IV composed of maximum tomato genotypes i.e. 88572, Gol, Nagina, Checo, Pant Bahar and Avinash. Third experiment deals with Agrobacterium mediated transformation of DREB 1A gene in tomato using tissue culture. Prior to transformation three tomato genotypes were subjected to tissue culture studies to select the best genotype for callus induction and regeneration. Hypocotyls and leaf discs were used as explant source and inoculated on Murashige and Skoog media having different hormonal combinations. The callus induction and regeneration frequency varied among genotypes and it was concluded that hypocotyls of tomato genotype Avinash produced maximum callus induction and regeneration frequency on MS media xiiisupplemented with IAA (0.5 mg/l) and Kin (1.5 mg/l). Agrobacterium mediated gene transformation was carried out in tomato genotype Avinash to transfer DREB 1A gene tolerant to salt stress. During the study it was observed that two min infection time with Agrobacterium, two days for co-cultivation, Acetosyringone @ 50 μM in co-cultivated media and 50 mg/l hygromycin in selection media was optimum for high frequency transformation. Polymerase chain reaction (PCR) analysis was conducted to confirm trasfomants by amplification of 500 bp size DNA fragment for hygromycin resistance gene.