Genetic Engineering of Pinus Radiata and Picea Abies, Production of Transgenic Plants and Gene Expression Studies

Walter, Christian; Bishop-Hurley, Sharon L.; Charity, Julia A.; Grace, Lynette J.; Höfig, Kai P.; Holland, L.; Möller, Ralf; Moody, Judy; Wagner, Annika; Walden, Adrian R.

doi:10.1016/s0921-0423(01)80075-7

Cited by 3 publications

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“…Its commercial importance has initiated strong efforts to better understand pine genetics (Walter et al 2001). In the area of reproductive development, male sterility was identified as a desirable trait for both scientific and commercial reasons.…”

Section: Introductionmentioning

confidence: 99%

Expression analysis of four Pinus radiata male cone promoters in the heterologous host Arabidopsis

Höfig

Moyle

Putterill

et al. 2003

Planta

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Four male cone-specific promoters were isolated from the genome of Pinus radiata D. Don, fused to the beta-glucuronidase (GUS) reporter gene and analysed in the heterologous host Arabidopsis thaliana (L.) Heynh. The temporal and spatial activities of the promoters PrCHS1, PrLTP2, PrMC2 and PrMALE1 during seven anther developmental stages are described in detail. The two promoters PrMC2 and PrMALE1 confer an identical GUS expression pattern on Arabidopsis anthers. DNA sequence analysis of the PrMC2 and PrMALE1 promoters revealed an 88% sequence identity over 276 bp and divergence further upstream (<40% sequence identity). GUS expression driven by a 276-bp PrMALE1 promoter fragment showed the same pattern in Arabidopsis anthers as observed for the full-length PrMALE1 promoter. Within the 276-bp promoter fragment a region of high homology to a previously described 16-bp anther-box was identified. In gain-of-function experiments the putative PrMALE1 anther-box was fused upstream of a 90-bp CaMV 35S minimal promoter, as a single copy in the sense direction and as an inverted repeat. No GUS expression was conferred to Arabidopsis anthers by either of these two constructs. In a loss-of-function experiment a 226-bp PrMALE1 deletion construct, which did not contain the putative PrMALE1 anther-box, still maintained the originally observed PrMALE1 GUS expression pattern. Hence, gain-of-function as well as loss-of-function experiments consistently showed that the putative anther-box of the PrMALE1 promoter is non-functional in the Arabidopsis genetic background. For the analysis of the four full-length pine promoters PrCHS1, PrLTP2, PrMC2 and PrMALE1, transformation vectors based on pCAMBIA2200 and pCAMBIA1302 were used. It will also be demonstrated in this article that sequences within the T-DNA borders of these vectors caused a characteristic histological background expression in Arabidopsis, with staining observed in vascular tissue of leaves, sepals, roots, filaments of stamens and in stems and pistils.

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Section: Introductionmentioning

confidence: 99%