Plant Tissue Culture for In Vitro Mutagenesis, Large-Scale Propagation, and Genetic Transformation

Abstract

An ever-increasing demand of uniform plants of commercially valuable plant species needs their clonal propagation on a large scale using different strategies of tissue culture. We have reported a large number of plants which were propagated in vitro using different plant parts. A number of problems were solved before large-scale propagation of these crops, like shoot organogenesis, multiplication of shoots, rooting, acclimatization and hardening, control of shoot browning/necrosis and defoliation, etc. Besides, popularization of temperate climate crops, e.g., Asiatic hybrid lilies and gerbera, to subtropical climate of Lucknow, was achieved using tissue culture. New varieties of chrysanthemum differing in color and shape of the florets were developed using in vitro mutagenesis.

Transformation protocols using Agrobacterium tumefaciens have been optimized in Jatropha curcas—a plant important for biofuel and Withania somnifera—an important Indian medicinal plant using leaf segments (LS). W. somnifera has some medicinally important sterol glycosyltransferases (SGT) which are the enzymes that glycosylate sterols and play an important role in providing tolerance to the plant against biotic and abiotic stresses. These genes were differentially expressed in different organs of the plant and also in response to biotic and abiotic stress. Functional characterization of WsSGT gene was done by its overexpression in homologous and heterologous expression systems of N. tabacum and A. thaliana, whereas for suppression/gene silencing, RNAi and artificial miRNA technologies were used. The transgenic plants showed improved germination and tolerance to salt, heat, and cold stress when compared to WT plants. Increased enzyme activity and sterol glycosides could be demonstrated through biochemical analysis. Stress-responsive elements were observed when the promoter of WsSGTL1 gene of W. somnifera was cloned and sequenced.