Plant tissue culture approach for cloning and conservation of some important RET medicinal plants

Abstract

In vitro protocol for rapid micropropagation/cloning in case of four medicinally important rare, endangered and threatened (RET) plants of Indo-Gangetic Plain, namely, Clerodendrum serratum, Uraria picta, Operculina petaloidea and Embelia tsjeriam-cottam, employing nodal stem segments of field-grown plant, was developed for the purpose of ex situ conservation. In all the four plants, sustained proliferation of shoots as well as 100 % induction of rooting in isolated shoots could be maintained even in long-term culture, and complete plantlets were produced, which grew luxuriantly and came to flowering under field conditions. The genetic fidelity of plants raised through tissue culture was assured by random amplified polymorphic DNA (RAPD) analysis in case of C. serratum, U. picta and E. tsjeriam-cottam. In U. picta, quantitative estimation of two isoflavonones, isolated from the roots of mother plant and from the tissue culture-raised plants, revealed no significant difference in their concentrations which further strengthened true-to-type nature of in vitro cloned plants. Another process was developed for rapid micropropagation of a medicinally important wild, endemic, primitive and endangered monoembryonic Citrus species, viz. C. indica, growing in thin population in Garo Hills in Northeastern Himalayan region of the country. Such in vitro developed processes may be utilized for the supply of enough raw materials to various pharmaceutical companies and for providing large number of cloned plants to re-establish them in their natural habitats for in situ conservation and sustainable utilization. In case of Azadirachta indica, a medicinally important tree, a process for in vitro clonal multiplication of a 40-year-old mature tree through nodal stem segments was standardized. Further multiplication rate was augmented by inducing differentiation of multiple shoots from leaflet segments excised from in vitro raised proliferating shoots. In order to preserve the germplasm of highly heterozygous elite neem trees, an innovative method of long-term regenerative excised root culture was developed. The genetic fidelity of field-grown plants raised through nodal stem segments of a 40-year-old mature tree and through root segments, taken from long-term excised root cultures, was ascertained by RAPD markers as well as through chemical analysis in respect of azadirachtin content.