Abstract
Zygotic embryos excised from immature green fruits of the rattan palm, Calamus thwaitesii and cultured for 16 weeks under optimum culture conditions in Murashige and Skoog (MS) medium supplemented with 31.67 μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 35.23 μM 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) produced mixed (compact and friable) calli at 70 and 92 % rates. The semi-friable part of the callus (~500 mg) separated and subcultured in medium containing 2.22 μM 6-benzyladenine and 1.07 μM α-naphthalene acetic acid produced groups of 10.37 ± 0.60–21.52 ± 0.48 discrete globular embryoids of varied size in 6–8 weeks. Calli raised in presence of 2,4,5-T were relatively more prolific, friable and embryogenic than those induced by 2,4-D. Embryoids (2.0–3.0 mm) isolated and cultured in basal medium germinated into plantlets at 65 % efficiency while the immature (0.5–2.0 mm) ones produced calloid structures. Approximately 15 % of the in vitro plantlets raised from the 2,4-D-induced embryogenic calli produced secondary immature embryoids on the sheath and lamina parts of leaves which were isolated and cultured in basal medium developed into rooted plantlets at 62 % rate in 12–16 weeks. The continued growth of the embryo-derived callus through successive subcultures together with differentiation of embryoids into plantlets, and the formation of immature embryoids on in vitro plantlets in MS basal nutrient medium reports for the first time a reliable method of producing at least 116 plants from a single embryo in a year. Rooted plantlets treated with 50 % glycerin survived at 78 % rate after hardening and 82.7 % of the hardened plants reintroduced into forest segments showed uniform growth free of morphological abnormalities after 3 years of observation. In addition to embryogenesis, cryopreservation of the zygotic embryos through simple drying and encapsulation–dehydration methods resulting 60–70 % recovery rates also offers another option for long-term conservation and sustainable utilization of this plant genetic resource.
Highlights
Rattans are unique and versatile group of spiny climbing palms or canes with solid stem distributed in the forests of Indo-Malayan and African regions
Zygotic embryos excised from immature green fruits of the rattan palm, Calamus thwaitesii and cultured for 16 weeks under optimum culture conditions in Murashige and Skoog (MS) medium supplemented with 31.67 lM 2,4-dichlorophenoxyacetic acid (2,4-D) and 35.23 lM 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) produced mixed calli at 70 and 92 % rates
While callusing followed by differentiation of shoots and roots was observed in 5–15 % of the embryos cultured even in high concentrations (31.67 lM) of 2,4-D (Fig. 1a), organogenesis was altogether arrested at concentrations exceeding 11.74 lM 2,4,5-T (Table 1)
Summary
Rattans are unique and versatile group of spiny climbing palms or canes with solid stem distributed in the forests of Indo-Malayan and African regions. Species of Calamus distributed in the Western Ghats region of southern peninsula are a high value non-timber forest product primarily used for making furniture, baskets and handicraft items and an important raw material of the cottage industry contributing employment to approximately 1,00,000 people in the state of Kerala alone (Bhat et al 1989). Due to shrinkage of wild rattan resources and non-availability of raw materials, the rural industrial units are either closed down or forced to receive their supplies from far away Assam and Andaman islands, causing significant increase in the prices of cane products and jeopardizing the livelihood of workers involved in the local extraction and processing of canes. The most seriously affected economically important species in Kerala State in India is Calamus thwaitesii which despite its wide distribution is indiscriminately exploited to the extent that mature useful canes are no more available in the accessible forests. Extraction of canes is often done before flowering and seed bearing mature palms are confined to protected forests in wildlife sanctuaries and national parks
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