Stressful environment is a limiting factor for rubber productivity in the traditional rubber growing areas and limits the expansion of rubber cultivation to newer areas in several rubber producing countries, including India. Conventional methods of Hevea breeding take several years to produce plants with desirable traits. Genetic transformation offers a viable approach for producing plants with desired traits within a short period of time. Since osmotin gene is reported to impart tolerance to abiotic and biotic stress, the present study on Agrobacterium mediated gene transfer was carried out with the gene encoding osmotin protein. Osmotin gene construct with CaMV 35S promoter and kanamycin as selectable marker was used for the purpose. Transformation frequency of 48 per cent was obtained from anther derived embryogenic calli. Molecular analysis with PCR and RT-PCR could confirm the integration of gene and its expression. The calli were subjected to PEG as well as salt stress for fixed intervals. Higher proline accumulation in tansgenic cell lines compared to control was observed under PEG stress. Transgenic cell lines could tolerate higher concentration (50 mM). The regenerated plants from these transgenic lines are expected to perform well under stressful environments.