Micropropagation of Hadrolaelia grandis through transverse and longitudinal thin cell layer culture

Abstract

An efficient in vitro propagation method was established for Hadrolaelia grandis using transverse and longitudinal thin cell layer (tTCL and lTCL, respectively) culture systems. Two and three-month old protocorms from in vitro germinated seeds were used for this study. tTCl and lTCL (0.5 mm thick) from protocorms were cultured on Woody Plant Medium (WPM) supplemented with benzyladenine (BA) (0, 1.1, 2.2, 4.4, 8.8 and 17.6 μM). The elongation and root development of protocorm-like bodies (PLBs) were evaluated in medium containing activated charcoal (0, 1, 2 and 3 g·L−1) or indole-3-butyric acid (IBA) (0, 1.25, 2.5, 5.0 and 10 μM). The plantlets were transplanted in polystyrene trays containing: coconut powder, pine bark with humus worm and vermiculite. The highest percentage (70.8%) and total number of PLBs per explant (30.1) were obtained from transversal protocorm sections two-month old cultured on WPM medium supplemented with 2.2 μM BA. The lTCL technique was more efficient with sections from three-month old protocorms grown in medium containing 8.8 μM BAP (83.3% of PLBs and 34.4 PLBs regenerated per explant). PLB development was optimal on WPM medium containing 1.0 g·L−1 activated charcoal, and IBA was not necessary for rooting. Plantlets were successfully acclimatized (with a survival rate of 98.2%) when vermiculite was used as a substrate.