B. Venkateswarlu

Metabolic engineering using mtlD gene enhances tolerance to water deficit and salinity in sorghum

Sorghum bicolor L. Moench cv. SPV462 was transformed with the mtlD gene encoding for mannitol-1-phosphate dehydrogenase from E. coli with an aim to enhance tolerance to water deficit and NaCl stress. Transgene (pCAM mtlD) integration and expression were successfully confirmed by PCR, Southern, RT-PCR and Western analysis. Segregation analysis based on germination of T0 seed on hygromycin-supplemented medium revealed an expected Mendelian ratio 3:1 in lines 5, 72 and 75. Retention of leaf water content was remarkably higher in transgenic leaf segments when exposed to polyethylene glycol 8000 (−2.0 MPa), as compared to the untransformed controls. Another significant finding is that the transgenics maintained a 1.7 to 2.8 fold higher shoot and root growth, respectively, under NaCl stress (200 mM) when compared to untransformed controls. These results demonstrate that engineering mannitol biosynthetic pathway into sorghum can impart enhanced tolerance to water deficit and salinity.

Efficient plant regeneration from shoot apices of sorghum

An efficient and rapid regeneration protocol was developed using shoot apices from germinating seedlings of two cultivars of sorghum, SPV-462 and M35-1, as explants. A vertical slit given from the base of each dissected apex enhanced the efficiency of callusing response by two fold. MS medium containing 0.5 mg dm−3 each of 2,4-D and kinetin was most effective in producing friable and embryogenic calli. Scanning electron microscopy of these calli detected somatic embryogenesis. Calli thus induced gave rise to approximately 42 green shoots per callus in both the genotypes when transferred to regeneration medium containing 1.5 mg dm−3 kinetin.

In vitro screening of Vigna mungo genotypes for PEG induced moisture deficit stress

Two black gram (Vigna mungo L. Hepper) genotypes LBG20 and PU19 were selected to study the impact of PEG induced drought stress on seed germination, metabolite concentration and activities of antioxidant enzymes. Stress caused considerable decrease in germination and fresh weight of seedlings of both the genotypes. It led to increase in protein concentration, contents of starch and total soluble sugars while decrease was observed in the activities of antioxidant enzymes, contents of free amino acids, reducing sugar and total phenols. SDS-PAGE analysis indicated accumulation of some proteins with the germination under stress conditions. LBG20 which showed increase in soluble sugars, starch, proteins and higher activities of antioxidant enzymes was observed to be relatively more tolerant to drought stress over PU19.