RESUMO
An efficient in vitro propagation method was established for Brasilidium forbesii (Hook.) Campacci using transverse and longitudinal thin cell layer (tTCL and lTCL, respectively) culture systems. Six-month-old protocorms from in vitro germinated seeds were used for this study. TCLs (1.0-mm thick) from protocorms were grown on Woody Plant Medium (WPM) supplemented with benzyladenine (BA) (0.54.0 µM).The lTCL technique was more efficient for inducing protocorm-like bodies (PLBs) and regenerating shoots than the tTCL technique. The frequency of PLB formation was influenced by BA concentration, and the lTCL explant grown on a medium containing 2.0 µM BA produced the highest percentage of new protocorms (77%) with a total of 22.7 PLBs per explant, after the first subculture on the same medium. Plantlet development was optimal on WPM medium containing 3.0 g L -1 activated charcoal, and indole-3 -butyric acid was not necessary for rooting. Regenerated plants were successfully acclimatized in a greenhouse after 16 weeks using vermiculite as the substrate (100% survival).
Foi estabelecido um método eficiente de propagação in vitro para Brasilidium forbesii (Hook.) Campacci utilizando a técnica 'thin cell layer' transversal (TCLt) e longitudinal (TCLl). Foram utilizados protocormos de seis meses obtidos da germinação in vitro. TCLs (1,0 mm de espessura) dos protocormos foram cultivados no meio 'Woody Plant Medium' (WPM), acrescido com benziladenina (BA) (0,5 a 4,0 µM). A técnica TCLl foi mais eficiente para indução de estruturas semelhantes a protocormos (ESPs) e regeneração de brotações do que a técnica TCLt. A frequência de formação de ESP foi influenciada pela concentração de BA e o explante TCLl, cultivado em um meio contendo 2,0 µM BA, produziu a mais alta percentagem de novos protocormos (77%), com um total de 22,7 PLBs por explante, após o primeiro subcultivo para o mesmo meio. O desenvolvimento das plântulas foi eficiente no meio WPM contendo 3,0 g L -1 de carvão ativado e o ácido indol-3- butírico (AIB) não foi necessário para o enraizamento. Plantas regeneradas foram estabelecidas com sucesso em casa de vegetação, utilizando vermiculita como substrato (100% de sobrevivência), após 16 semanas.
Assuntos
Técnicas In Vitro/métodos , OrchidaceaeRESUMO
Objective To screen the optimal medium formula for obtaining high proliferation rate and differentiation rate of protocorm-like body ( PLB) of Dendrobium officinale Kimura et Migo ( D.officinale) . Methods We observed the effect of different concentrations of phytohormones such as 6-benzylaminopurine (6-BA) , 1-naphthaleneacetic acid ( NAA) , 2,4-dichlorophenoxyacetic acid ( 2,4-D) and kinetin ( KT) , or their combinations together with organic additives such as potato extractive ( PE) , banana extractive ( BE) , apple extractive ( AE) and coconut milk ( CM) on the proliferation and differentiation of PLB of Dendrobium officinale Kimura et Migo. Results 6-BA, NAA and KT at certain concentrations were beneficial to the proliferation and growth of PLB, and when the concentration of KT was 1.0 mg/L, 50-d PLB proliferation times reached to 7.20. 2,4-D had obvious inhibitory effect on PLB proliferation and growth. The combination of 6-BA, NAA and KT showed stronger effect on PLB proliferation than the phytohormones used alone, and the combination of 0.5 mg/L 6-BA, 1.0 mg/L KT and 1.0 mg/L NAA had the best effect on PLB proliferation, 50-d PLB proliferation times reaching to 9.52. Certain concentrations of PE and AE could promote PLB differentiation, and 100 g/L PE brought well-grown seedling in test tube and the highest differentiation rate, being up to 92.6% . Conclusion The optimal hormone matching and additives concentration on proliferation and differentiation of PLB of Dendrobium officinale Kimura et Migo have been obtained, which can lay a foundation for its rapid propagation and artificial seed preparation.