An efficient procedure to stably introduce genes into an economically important pulp tree (Eucalyptus grandis x Eucalyptus urophylla)

• Published in: Transgenic research Vol. 12; no. 4; pp. 403 - 411
• Main Authors: TOURNIER, Vincent, GRAT, Sabine, MARQUE, Christiane, EL KAYAL, Walid, PENCHEL, Ricardo, DE ANDRADE, Gisele, BOUDET, Alain-Michel, TEULIERES, Chantal
• Format: Journal Article
• Language: English
• Published: Dordrecht Springer 01.08.2003; Springer Nature B.V
• Subjects: Base Sequence; Biological and medical sciences; DNA Primers; Eucalyptus - genetics; Eucalyptus grandis; Eucalyptus gunnii; Eucalyptus urophylla; Fundamental and applied biological sciences. Psychology; Gene Transfer Techniques; cinnamyl alcohol dehydrogenase; juvenile clones; genetic engineering; eucalyptus; antisense
• ISSN: 0962-8819; 1573-9368
• DOI: 10.1023/A:1024217910354

Regeneration problems are one of the main limitations preventing the wider application of genetic engineering strategies to the genus Eucalyptus. Seedlings from Eucalyptus grandis x Eucalyptus urophylla were selected according to their regeneration (adventitious organogenesis) and transformation capacity. After in vitro cloning, the best genotype of 250 tested was transformed via Agrobacterium tumefaciens. A cinnamyl alcohol dehydrogenase (CAD) antisense cDNA from Eucalyptus gunnii was transferred, under the control of the 35S CaMV promoter with a double enhancer sequence, into a selected genotype. According to kanamycin resistance and PCR verification, 120 transformants were generated. 58% were significantly inhibited for CAD activity, and nine exhibited the highest down-regulation, ranging from 69 to 78% (22% residual activity). Southern blot hybridisation showed a low transgene copy number, ranging from 1 to 4, depending on the transgenic line. Northern analyses on the 5-16 and 3-23 lines (respectively one and two insertion sites) demonstrated the antisense origin of CAD gene inhibition. With respectively 26 and 22% of residual CAD activity, these two lines were considered as the most interesting and transferred to the greenhouse for further analyses.