Abstract

Gentiana straminea Maxim is an endangered medicinal plant in the Qinghai-Tibet Plateau. To speed up the production of this species, an in vitro protocol for efficient plant regeneration was developed from its leaf explants. Auxins and cytokinin alone or in combination were examined for their effects on callus induction and plant regeneration. The results indicated that Murashige and Skoog medium supplemented with 2 mg l^-1 2,4-dichlorophenoxyacetic acid and 0.5 mg l^-1 N6-benzylaminopurine (BA) was the best medium for embryogenic callus induction, while 3 mg l^-1 BA induced the highest frequency (93.8%) of regeneration and development of shoots. Regenerated plantlets with well-developed root systems were hardened in the greenhouse and successfully established in the soil. Genetic stability of the regenerated plants was assessed by 25 inter simple sequence repeat (ISSR) markers. Out of 25 ISSR markers, 14 produced clear, reproducible bands with a mean of 6.9 bands per marker. The results confirmed that the regenerants maintained high genetic fidelity.

Key words: Gentiana straminea, somatic embryogenesis, leaf explant, genetic stability, inter simple sequence repeat (ISSR).

Abbreviation

AFLP, Amplified fragment length polymorphism; BA, N6-benzylaminopurine; CTAB, cetyltrimethylammonium bromide; dNTPs,deoxyribonucleotide triphosphate; 2,4-D,   2,4-dichlorophenoxyacetic acid; ISSR,inter simple sequence repeat; MS, Murashige and Skoog; NAA,   α-naphthaleneacetic acid; PCR, polymarase chain reaction; RAPD, randomly amplified polymorphic DNA; RFLP, restriction fragment length polymorphism; UBC,University of British Columbia.