Medicinal plants are the most important source of life saving drugs and uses as a major source of medicine since thousand years ago. Using approach such as cell, organ and tissue culture, as well as genetic engineering and molecular markers, biotechnology is able to extend application and effacing of medicinal plant as renewable sources in production of medicine. Cell, tissue and organ culture, has facilitated the massive and rapid propagation of a large number of medicine plant. Propagated plants through in vitro methods are free-disease and uniform in terms of quality and genetics Cryopreservation is long-term conservation method in liquid nitrogen and provides an opportunity for conservation of endangered medicinal plants. In-vitro production of secondary metabolites in plant cell suspension cultures has been reported from various medicinal plants. Bioreactors are the key step towards commercial production of secondary metabolites by plant biotechnology. Genetic transformation may be a powerful tool for enhancing the productivity of novel secondary metabolites; especially by Agrobacterium rhizogenes induced hairy roots. This article discusses the applications of biotechnology for regeneration and genetic transformation for enhancement of secondary metabolite production in vitro from medicinal plants.