Optimization of maturation process for somatic embryo production and cryopreservation of embryogenic tissue in Pinus koraiensis

Abstract

Korean pine broadleaf mixed forests are important for maintaining biodiversity in Northeast China. Excessive harvesting of the Korean pine tree has led to it being listed as an endangered species in China. Somatic embryogenesis technology has the potential to accelerate breeding by improving seedling quality to meet future wood harvesting needs. In addition, rare and valuable germplasm resources can be maintained through the use of cryopreservation. Here, we described the utilization of somatic embryogenesis in Korean pine, with a focus on maturation and cryopreservation. Embryogenic tissues (ETs) were induced from immature seeds collected from open-pollinated sources of Korean pine (28 families). Highest induction percentage of ET reached 22%. The proliferation ability of ET in different cell lines differed. Increasing ABA(up to 80 µM L− 1), gellan gum (up to 12 g L− 1) and sucrose (0.2 m L− 1) concentrations in maturation medium improved the yield of cotyledonary somatic embryo (SE). Maximum yield of cotyledonary SEs (160 SEs g− 1 fresh mass) was obtained following ET dispersion on filter paper discs at low density (50 mg fresh mass). Both the germination percentage of SEs and survival percentage of regenerated plants were about 50%. Moreover, ET began to lose their potential for somatic embryogenesis after 6 months of subculture. ET cultures were cryopreserved, and culture recovery (133 SEs g− 1 fresh mass) was demonstrated after thawing. In this paper we report a significantly improved procedure for generating Korean pine SEs, leading to a high yield of cotyledonary SEs. A cryopreservation protocol for the ET of Korean pine was also established. The present work reports a significantly improved procedure for generating somatic embryos and cryopreservation of embryogenic tissue in Korean pine.