Tissue and cell-specific expression of eucalyptus CAD promoter in transgenic poplar

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Feuillet, Catherine | Pilate, Gilles | Boudet, A.M. | Grima Pettenati, Jacqueline

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*INRA, Station d'Amélioration des Arbres Forestiers, Orléans Diffusion du document : INRA, Station d'Amélioration des Arbres Forestiers, Orléans. International audience. Cinnamyl alcohol dehydrogenase (CAD) catalyses the last step in the biosynthesis of the lignin monomers. We recently reported the isolation of a CAD cDNA from Eucalyptus gunnii and showed that the CAD mRNA was most abundant in xylem, providing supporting evidence for the role of CAD in lignification [Grima-Pettenati et al, 1993, PMB, 21, 1085-1095]. We have now isolated the corresponding CAD genomic clone (geucad 16). Sequence comparison with the CAD CDNA allowed the identification of four introns. A 2.5 kB promoter fragment was transcriptionnally fused to the B-glucuronidase (GUS) gene and shown to be functionally expressed in Eucalyptus protoplasts. In these transient gene expression experiments, the CAD promoter is at least twice as high as the 35S CaMV promoter. To study the developmental and spatial regulation of the CAD gene, the chimeric CAD-GUS was introduced into poplar (Populus tremuloides x Populus alba) via Agrobacterium tumefaciens transformation, Eight independant transgenic plants were analyzed in detail. Quantitative fluorimetric analysis revealed that GUS activity was highest in roots followed by stems and leaves. Histochemical staining using X-gluc indicates a specific expression in the vascular tissues of stems, roots, petioles and leaves. Staining stem sections for both GUS activity and lignin using phloroglucinol demonstrated that the CAD promoter was specifically expressed in cells surrounding the lignified phloem fibers and in the ray cells of the developing xylem. Analysis of 5' deletions of the CAD promoter are currently underway.

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