Medicinal plants are valuable sources of medicinal and many other pharmaceutical products. The conventional propagation method is the principal means of propagation and takes a long time for multiplication because of a low rate of fruit set, and/or poor germination and also sometimes clonal uniformity is not maintained through seeds. The plants used in the phyto-pharmaceutical preparations are obtained mainly from the natural growing areas. With the increase in the demand for the crude drugs, the plants are being overexploited, threatening the survival of many rare species. Also, many medicinal plant species are disappearing at an alarming rate due to rapid agricultural and urban development, uncontrolled deforestation, and indiscriminate collection. Advanced biotechnological methods of culturing plant cells and tissues should provide new means for conserving and rapidly propagating valuable, rare, and endangered medicinal plants. The present investigations were carried out with a view to standardize an in-vitro culture technique for propagation of Tinospora cordifolia Hook. The Nodal and inter nodal segments from healthy grown plants were used as explants. For culturing Explants were cultured on standard Murashige and Skoog (MS) medium supplemented with different concentrations of benzyl amino purine (BA), kinetin (Kn) and adenine (Ad) for primary shoot proliferation. The shoot proliferation (50 % single shoot,) was observed in MS medium containing BA 5.0 + Kn 1.0.For rooting of the micro shoots, half strength MS medium supplemented with 0.4mg/L Naphthalene acetic acid (NAA) exhibited best results with average rooting response of 40 %. After acclimatization and transplantation, 100% of the in- vitro derived plants were found healthy in ex vivo conditions.