Somatic embryogenesis from immature and mature zygotic embryos, and embryo regeneration from protoplasts in black spruce ( Picea mariana Mill.)

Abstract

Embryogenic calli of black spruce ( Picea mariana Mill.) were initiated in vitro from immature zygotic embryos collected during the maturation period, and with mature zygotic embryos dissected from seed which had been stored for 13 years. With immature zygotic embryos as explants, highest frequency of embryogenic calli (65%) was obtained using full strength von Arnold and Eriksson (LP) medium in the absence of casein hydrolysate (CH). With mature zygotic embryos, embryogenic calli developed in 8% of explants on this medium and in 10% of explants using 1 2 strength modified Litvay ( 1 2 LM ) medium. Calli derived from mature embryos were grown as suspensions in liquid medium. Protoplasts isolated from six of these suspension lines differed in their response to culture conditions. In 1 2 LM (liquid, or agarose solidified) with 0.37 M glucose as osmoticum, protoplasts of two lines showed first divisions after 24 h and recognizable somatic embryos had developed by 7 days. Protoplasts from the other lines either failed during their development into somatic embryos, or developed into green non-embryogenic callus.