Efficient Method for Direct Embryogenesis in Phalaenopsis Orchid

Abstract

Light spectrum is one of the environmental cues that influence plant growth and development. Light is a stimulating factor for induction of somatic embryos during tissue culture practices. To accelerate the direct embryogenesis, six different light spectra including: white (W), red (R), blue (B), green (G), red + blue (R+B) and red + far red (R+FR) together with dark condition (D), in combination with thidiazuron (TDZ) in four concentrations (0, 0.5, 1.5 and 3 mg L-1) were used. Inter-simple sequence repeat was used for identification and genetic stability analysis of somatic regenerated plantlets. Intact protocorm explants showed higher potential for direct somatic embryogenesis (DSE) than the other explants. The rate of DSE was highly dependent on the concentration of TDZ and its interaction with light spectra. R and R + FR spectra with 3 mg L-1 TDZ on intact protocorms and R+FR with 3 mg L-1 TDZ were efficient treatments to induce DSE without somaclonal variation. G light spectrum has also significant effects on DSE of protocorm explants. The amplified products showed 26 scorable bands and regenerates were completely identical to the mother plant. In conclusion, this protocol provides way to regenerate plants through embryogenesis, and is a reliable protocol to obtain proper development and genetic stable Phalaenopsis embryos.