Efficient somatic embryogenesis and regeneration from leaf main vein and petiole of Actinidia chinensis planch. via thin cell layer culture technology

Abstract

Somatic embryogenesis (SEMs) is an effective micropropagation method that has been successfully applied to many different plant species; however, little is known about the SE in Actinidia chinensis Planch. - A species with high economic value. In this study, the efficiency of SEMs from leaf main vein and petiole of A. chinensis (seedlings germinated from seeds sterilized with 200 ppm silver nanoparticles for 7 min) via thin cell layer (TCL) technique was performed and compared with that of whole leaf main vein and whole petiole explants. The leaf main vein (mv - 1 mm × 10 mm in size) and leaf petiole (p - 1 mm × 10 mm in size) explants of in vitro A. chinensis were cut through transverse and longitudinal TCL (mv-tTCL and mv-lTCL for mv explants or p-tTCL and p-lTCL for p explants) with different sizes (½, ¼, 1/6, 1/8 of mv-lTCL or p-lTCL and ½, 1/3, ¼ of mv-tTCL or p-tTCL, respectively). Somatic embryo (SE) was recorded in the wounds of mv and p explants after 8 weeks of culture. For mv explants which cultured on MS medium supplemented with 0.02 mg/L NAA, 0.5 mg/L TDZ, 30 g/L sucrose and 8 g/L agar – MSE medium, the ½ mv-tTCL explant (mv was cut into 2 tTCL explants) gave the highest SEMs (98.67%) and number of somatic embryos per explant (10.66 embryos); meanwhile, p explants cultured on MSE medium, the ½ p-tTCL explant (p was cut into 2 tTCL explants) obtained the highest number of somatic embryos per explant (8.66 embryos) after 8 weeks of culture. In addition, the growth correction factor of SEMs was optimal at ½ mv-tTCL explant (21.04) or 1/6 p-lTCL (p was cut into 6 lTCL explants) and 1/8 p-lTCL (p was cut into 8 lTCL explants) (31.36 and 32.00). The current study has shown that the procedure of leaf main vein and petiole TCL derived- SEMs of A. chinensis is the optimal one. Large amounts of somatic embryo induction were observed from mv-TCL and p-TCL with full shapes of the globular, heart, torpedo and cotyledon. The shoot regeneration from somatic embryo with the cotyledon shape was optimal on medium containing 1.0 mg/L BA in terms of total number of shoots, shoot height and number of shoots. In addition, plantlets derived from shoot cultured on the MS medium supplemented with 0.4 mg/L NAA and planted in plastic pots with the combination of humus, coconut fiber and biofertilizers substrates (with ratio of 2:3:1 in weight) showed higher acclimatization and growth than those from other treatments in the greenhouse.