STABLE GENETIC-TRANSFORMATION OF PICEA-MARIANA (BLACK SPRUCE) VIA PARTICLE BOMBARDMENT

Stable genetic transformation of Picea mariana (black spruce) was obta ined via particle bombardment into two target tissues, mature cotyledo nary somatic embryos and suspensions from embryonal masses, with the B iolistic PDS-1000/He device. Seven transgenic embryogenic cell lines w ere obtained from the mature cotyledonary somatic embryos after second ary somatic embryogenesis from two different cell lines (R4F14 and 119 794-014). The suspension cultures from embryonal masses produced five transgenic cell lines from one cell line (R4F14). Expression of the in troduced beta-glucuronidase (GUS) and neomycin phosphotransferase II ( NPT II) genes was detected by histochemistry and fluorometry, and by E LISA in 10 of the lines. Two lines showed only NPT II gene expression. Four of the five lines obtained after bombardment of suspensions of e mbryonal masses showed lower levels of expression of GUS and NPT II. T he integration of the foreign genes was confirmed by polymerase chain reaction analyses and Southern hybridization for GUS and NPT II, and c omplex hybridization patterns were observed. The 12 transgenic lines o btained had a typical embryogenic morphology and were capable of matur ation and germination. Over 40 transgenic trees were regenerated from one of the transgenic lines, and they have a normal phenotype.