Comparative studies of protoplast development in jack pine (Pinus banksiana)

Abstract

Protoplasts were isolated either from the cotyledons of 8- to 10-day-old seedlings, or from cell suspensions, initiated from seedlings of the same open-pollinated seedlot of jack pine (Pinus banksiana Lamb.). Protoplast yields were 2.0 – 7.0 × 105/g fresh wt. for cotyledon-derived, and 0.3–5 × 105/g fresh wt. for suspension-derived protoplasts. Cotyledon protoplasts (6.8%) divided after 5 to 8 days incubation in liquid Lainé medium C with 450 mM glucose as osmoticum. Small clusters of 6 to 10 cells, which were observed after approximately 2 weeks, formed microcalli upon further dilution of the culture medium. By 6 to 8 weeks, these microcalli could be transferred to solid medium using a nurse culture of jack pine cells. Protoplasts isolated from seven cell suspension lines differed in their responses to culture conditions. First divisions were only observed with two cell lines cultured in either Lainé medium C or one-half Litvay with 370 mM glucose, but this did not lead to callus development. Key words: jack pine, Pinus banksiana, cotyledons, protoplasts, regeneration, callus.