EXPRESSION OF GUS IN SOMATIC EMBRYO CULTURES OF BLACK SPRUCE AFTER MICROPROJECTILE BOMBARDMENT

Immature embryos (stage I) and cotyledonary somatic embryos (stage III) of black spruce [Picea mariana (Mill) B.S.P.] were bombarded with tungsten particles coated with a gene construct containing the fusion of gus::nptII. GUS (beta-glucuronidase) activity was monitored histochemically with X-gluc giving a blue colour where transient gene expression was detected in the bombarded tissues. A high transient expression of gus was observed in stage I embryo cultures 2 d after bombardment (202 GUS foci per 300 mg tissue). GUS activity had substantially diminished in this material 14 d after bombardment, when grown in liquid LP maintenance medium containing BA (4.4 mu M), 2,4-D (9 mu M) and 1% sucrose. However, when stage I embryos were cultured on LP maturation medium containing ABA (40 mu M), IBA (1 mu M), 3.4% sucrose and 0.8% agar, GUS activity after 2 d was 335 GUS foci per 300 mg tissue, and the activity was detected until 30 d after bombardment. With stage III somatic embryos cultured on LP maintenance medium, 92% showed GUS activity 2 d after bombardment (16 GUS foci per embryo), and 31% showed activity 30 d after bombardment (4 GUS foci per embryo). GUS activity was still evident in 12% of the embryos (2 GUS foci per embryo) 45 d after bombardment.