Non-Destructive Assessment of Growth Performance of Embryogenic Suspension Culture of Pinus kesiya (Royle ex. Gord.) in Shake-Flask and Self-Designed Bubble Bioreactor and Successful Regeneration of Plantlets from the Culture Systems

Abstract

Embryogenic suspensions of Pinus kesiya (Khasi pine) derived from embryogenic cultures obtained from immature zygotic embryos (intact megagametophytes) of Pk-04 genotype were grown in shake-flasks and self-designed bubble bioreactors. Conifer somatic embryogenesis could find its utility for rapid mass propagation if grown as suspension cultures. To minimize the destruction of embryogenic cultures during study, a few growth characteristics viz., sedimented cell volume and number of stage-I (early stage) and -III (cotyledonary) embryo formation in suspensions were determined over a 21-day period. Compared to shake-flasks, proliferation in bioreactors resulted in increased biomass. The suspension culture showed a mixture of freely suspended as well as aggregates of stage-I (early stage) embryos. Maturation of somatic embryos was done on gelled medium and in submerged culture where gelled solid medium was covered with a layer of liquid medium. The number of stage-III (cotyledonary) embryos formed in bubble bioreactor were significantly higher than shake-flask cultures with both solid medium and submerged cultures. The cotyledonary embryos germinated successfully producing plantlets on germination medium.