Seed germination and tissue culture of Cymbidium giganteum Wall. ex Lindl.

Efficient protocols were established for in vitro seed germination, neo-formation of secondary (2) protocorms from primary (1 degrees) protocorms and multiple shoot buds and protocorm-like body (PLB) induction from pseudo-stem segments of in vitro-raised seedlings of Cymibidium giganteum. Four nutrient media, namely Murashige and Skoog (MS), Phytamax (PM), Mitra et al. (M), and Knudson 'C' (KC) were evaluated for seed germination and early protocorm development. In addition, the effects of peptone, activated charcoal (AC) and two plant growth regulators [6-benzylaminopurine (BAP) and 2,4-dichlorophenoxyacetic acid (2,4-D)] were also studied. Both M and PM supplemented with 2.0 g l(-1) peptone or 1.0 mg l(-1) BAP resulted in similar to 100% seed germination. Media supplemented with 2.0 g l(-1) AC could effectively induce large protocorms (1.6 +/- 0.1 mm in diameter). Neo-formation of 2 protocorms from 1 degrees protocorms was achieved in liquid and agar-solidified PM medium fortified with different concentrations and combinations of auxins (alpha-naphthalene acetic acid (NAA) and 2,4-D) and cytokinins [BAP and kinetin (KN)]. The highest number of 2 degrees protocorms was obtained in liquid medium (10.7 +/- 0.9/1 protocorm) supplemented with 2.0 mg l(-1) BAP + 1.0 mg l(-1) NAA. Although protocorms proliferated profusely in liquid medium, these did not develop further unless transferred to agar-solidified medium within 6-8 weeks, Multiple shoot buds and PLBs were induced from pseudo-stern segments on agar-solidified PM medium fortified with different concentrations and combinations of BAP and NAA and the maximum number of PLBs (6.00 +/- 0.20) was recorded when BAP and NAA were applied at 2.0 mg l(-1) each. A solid root system was induced from PLBs and shoot buds when these were transferred to half-strength PM or M media fortified with 0.5 mg l(-1) indole-3-acetic acid. Well-rooted plants were transferred to the greenhouse with 95% survival. (C) 2009 Elsevier B.V. All rights reserved.