PLANT CELL BIOTECHNOLOGY AND MOLECULAR BIOLOGY

Arnebia hispidissima (Lehm.) A.DC. (Boraginaceae), a red dye-yielding and medicinally important plant, is commonly distributed in arid/semi-arid regions of Rajasthan. This paper highlights an in vitro callus proliferation protocol for production of shikonin and its derivatives. Immature seeds explants were found to be most favorable for callus induction (74.6±1.52%). During callus induction, the maximum percentage response (78.9±1.57) was achieved on Murashige and Skoog (MS, 1962)  medium with additives supplemented with NAA (1.0 mg L^-1) and 2, 4-D (2.0 mg L^-1). Dye producing red colored globular and fast-growing (four-fold) callus was achieved on 2, 4-D (0.1 mg L^-1), Kin (0.5 mg L^-1) and BAP (0.1 mg L^-1). In vivo root extract and in vitro proliferated callus extract were compared using thin layer chromatography (TLC). Six and four promising bands were recorded from in vivo root extract and in vitro callus extract, respectively on the Methanol : Chloroform (1:9) extraction system. Three different types of calli (red, dark red and green) growing on varying media composition were studied for differential gene expression using SDS – PAGE which revealed presence of seven, twelve and twelve protein bands, respectively. The presence of one extra band (21.4 kDa) in dark red callus might be responsible for dye/shikonin derivative production. The results obtained in this study are preliminary and requires further analysis of protein profile/proteome in order to understand the shikonin production mechanism in this plant.