Breeding for crop resistance to abiotic stress is difficult due to its multigenic nature. An alternative approach is through DNA mutation and selection. An efficient in-vitro screening method has been developed for cauliflower to create mutant lines for resistance to salinity, hydroxyproline and frost. Mutant lines (derived NEU and NMU) and Control plants were subcultured several times on S23 media and then tested for salt, hydroxyproline resistance in-vitro and in-vivo. Leaf discs and leaf strips were plated on to solid media S23 with two concentrations of hydroxyproline (3 and 10 mM) and NaCl (350 and 550 mM) in order to study the resistance of hyp and NaCl of in-vitro and in-vivo plants, respectively. Shoot buds or meristems were exposed to 3 mM of hyp and 350 Mm NaCl to study their effect on proline content in leaves after 4 weeks. Shoots were grown in-vitro and weaned to in-vivo conditions. After 60 days from weaning the plantlets were assessed for frost resistance under acclimation (2 oC for 14 days) and non-acclimation conditions. Frost treatments at -3, -5 and -7 oC were applied. Assays showed that control plants had no salt or hydroxyproline resistance and selected plants had varying degrees of resistance for both in-vitro and in-vivo plants. Leaf proline content increased markedly in the mutant lines and the greatest proline contents occurred following NaCl stress with the most respondent line having 100 fold levels compared to the controls. Freezing injury was found to decrease with cold hardening and the mutant plants showed a greater capacity to acclimate than control lines. The results clearly demonstrated that proline accumulation could be used as a biochemical marker for discrimination between genotypes under abiotic stress. It was clearly shown that NaCl and hyp resistance were stable traits maintained for more than 15 in-vitro subcultures.