Plant growth analysis
Total germination was expressed as a percentage of the control for each variety. The germination percentage, root
length, shoot length, and root/shoot ratio of seedling were recorded on the 21st day of germination.
Biochemical analyses
Determination of proline content
Free proline content was estimated by following the method of Bates et al. (1973). Fresh 0.5 gm root and shoot
samples were homogenized in 5 ml of 3% sulpho salicylic acid using a mortar and pestle. About 2 ml of extract was
taken in test tube and to it 2 ml of glacial acetic acid and 2 ml of ninhydrin reagent were added. The reaction mixture
was boiled in a water bath at 100°C for 30 min. After cooling the reaction mixture, 4 ml of toluene was added. After
thorough mixing, the chromophore containing toluene was separated and absorbance of red color developed was
read at 520 nm against toluene black on UV-visible spectrophotometer (Chemito, UV- 2600). The proline
concentration was determined using calibration curve and expressed as mg proline per g fresh weight of tissue