Induction of somatic embryogenesis and plant regeneration in the reed grass (Phragmites communis Trin.)

Abstract

An in vitro culture system for the large-scale propagation of Phragmites communis Trin. (reed) was established by optimizing culture conditions for callus induction and differentiation together with plant propagation using regenerated plantlets. Callus was induced from stem segments with callus induction medium containing auxin: 4-fluorophenoxyacetic acid (4-FA) or 2,4-dichlorophenoxy acetic acid (2,4-D). A high frequency of callus induction was observed at relatively low concentrations (0.5 and 1 mg L^-1) of both 2,4-D or 4-FA. However, high concentrations (3, 4 and 5 mg L^-1) of either auxin suppressed callus induction. When applied for the first time, 1.0 mg L^-1 4-FA markedly improved the frequency of callus induction (up to 93%). The callus was then transferred to MS medium supplemented with 1.0 mg L^-1 2,4-D to promote the formation of embryogenic calli. The calli were grown in MS supplemented with different concentrations of 2,4-D and naphthaleneacetic acid (NAA) alone or in combination with benzyladenine (BA). After seven weeks of culture, the regeneration efficiency was determined for the calli maintained for the 45 differentiation media formulations. The highest regeneration capacity was obtained from the medium containing 0.05 mg L^-1 NAA and 2 mg L^-1 BA, and the combination of 0.2 mg L^-1 NAA and 2 mg L^-1 BA. Propagation of the regenerated plantlets was also examined in medium containing different sucrose concentrations; this experiment found that 60 g L^-1 sucrose showed the best growth rate. These improved regeneration and propagation systems could be used for bioreactor-based mass propagation or an in vitro culture system, and would be useful for transformation in Phragmites communis Trin.

Key words: Phragmites communis Trin, callus, regeneration, propagation, in vitro culture, 2,4-dichlorophenoxy acetic acid, 4-fluorophenoxyacetic acid.