Micropropagation of the Thai orchid Grammatophyllum speciosum blume
Protocorm-like bodies (PLBs) were induced from shoot tips of Grammatophyllum speciosum, a Thai orchid. The highest frequency of PLBs (93%) were observed on explants incubated on 1/2-Murashige and Skoog (MS) liquid medium containing 2% (w/v) sucrose without any plant growth regulators (PGRs). Tests with different carbon sources compared to sucrose revealed that maltose promoted the highest relative growth of G. speciosum PLBs (7-fold increase), while trehalose and sucrose yielded 5-fold and 4-fold increases, respectively. In 1/2 MS liquid medium, addition of 15 mg/l of chitosan promoted a 7-fold increase in PLB growth while 25 mg/l promoted a 4-fold increase. However, the relative growth rate in solid culture was significantly lower than that in liquid culture. In addition, chitosan supplementation in solid medium promoted shoot formation but not rooting. Plantlet regeneration was induced using a combination of NAA and BA supplementation in 1/2 MS solid medium with optimum induction shoot and root formation at 2.0 mg/l NAA and 1.0 mg/l BA. Using this protocol, approximately 8 months was required to obtain a hundred plantlets from one shoot tip. The plantlets showed no changes in ploidy when tested by flow cytometry.
KeywordsGrammatophyllum speciosum Micropropagation Chitosan Flow cytometry Protocorm-like bodies
This work was supported by a grant from The Royal Golden Jubilee Ph.D. Program, Thailand Research Fund, Thailand. We thank Professor T.W. Flegel for reviewing the manuscript. We are also grateful to Prof. Masahiro Mii, Chiba University, Dr. Pranee Fucharoen, Dr. M.L. Saovaros Svasti and Mr. Teerasakdi Chaiya, Thalassemia Research Center, Institute of Molecular Biosciences, Mahidol University, for kindly providing help in flow cytometry analysis and to the Japan Horticultural Production and Research Institute, Mastudo, Chiba for training.
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