Light strongly promotes gene transfer from Agrobacterium tumefaciens to plant cells

Abstract Light conditions during Agrobacterium-based plant transformation, the most routinely used method in plant genetic engineering, differ widely and, to our knowledge, have not been studied systematically in relation to transformation efficiency. Here, light effects were examined in two already optimized transformation procedures: coculture of Agrobacterium tumefaciens with callus from two genotypes of the crop plant Phaseolus acutifolius (tepary bean) and coculture of root segments from two ecotypes of Arabidopsis thaliana. Except for the light conditions during coculture, all steps followed established procedures. Coculture was done either under continuous darkness, under a commonly used photoperiod of 16 h light/8 h darkness or under continuous light. b-glucuronidase (GUS) production due to the transient expression of an intron-containing uidA gene in the binary vector was used to evaluate T-DNA transfer. In all situations, uidA expression correlated highly and positively with the light period used during coculture; it was inhibited severely by darkness and enhanced more under continuous light than under a 16 h light/8 h dark photoperiod. The promotive effect of light was observed with Agrobacterium strains harboring either a nopaline-, an octopine-or an agropine/succinamopine-type nononcogenic helper Ti plasmid. The observed positive effect of light has obvious implications for developing and improving transient and stable transformation protocols, specifically those involving dark coculture conditions

Overexpression of arginase in Arabidopsis thaliana influences defence responses against Botrytis cinerea

Arabidopsis possesses two arginase-encoding genes, ARGAH1 and ARGAH2, catalysing the catabolism of arginine into ornithine and urea. Arginine and ornithine are both precursors for polyamine biosynthetic pathways. We observed an accumulation of ARGAH2 mRNA in Arabidopsis upon inoculation with the necrotrophic pathogen Botrytis cinerea. Transgenic lines displaying either overexpression of ARGAH2 or simultaneous silencing of both Arabidopsis arginase-encoding genes were created and their resistance to B. cinerea infection evaluated. Overexpression of arginase resulted in changes in amino acid accumulation, while polyamine levels remained largely unaffected. Silencing lines were affected in both amino acid and putrescine accumulation. Arabidopsis plants overexpressing the arginase gene were less susceptible to B. cinerea, whereas silencing lines remained as susceptible as the wild type. We discuss how arginase might interact with plant defence mechanisms. These results provide new insights into amino acid metabolic changes under stress

An 11-Year Overview of the Belgian Donor and Transplant Statistics Based on a Consecutive Yearly Data Follow-up and Comparing Two Periods: 1997 to 2005 Versus 2006 to 2007

Background: The Belgian Transplant Coordinators Section is responsible for the yearly data follow-up concerning donor and transplantation statistics in Belgium and presents herein a 10-year overview. Methods: The procurement and transplant statistics were compared between 2 periods: Period 1 (P1, 1997-2005) versus Period 2 (P2, 2006-2007). Results: The kidney and liver waiting lists (P1 vs P2) showed an overall decrease for a period of 2 consecutive years in P2; kidney (-170 patients; -18%), and liver (-83 patients; -34%). All other waiting lists (heart, lung, pancreas) remained stable. Mean ED further increased (P1 vs P2); 229 (P1) versus 280 (P2, +22.27%). Non-heart-beating donors were significantly (+288%) more often procured in P2. Mean donor age was 37.9 ± 17.8 years (P1) versus 46.5 ± 19.9 years (P2), and mean organ yield per donor was 3.48 ± 1.7 (P1) versus 3.38 ± 1.8 (P2). Overall transplant activity per million inhabitants increased 21.1%. Conclusion: For 2 consecutive years, the Belgian statistics showed significantly increased donor activity with an impact on waiting list dynamics and transplantation. The mean organ yield per donor was not influenced despite an increased average age and change in reason for death. © 2009.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

Overview of the Belgian Donor and Transplant Statistics 2006: Results of Consecutive Yearly Data Follow-up by the Belgian Section of Transplant Coordinators

Background: The Belgian Section of Transplant Coordinators, created in 1997 under the auspices of the Belgian Transplant Society, is in charge of the collection of the national data about donor/procurement activities. Methods: Data are collected in all Belgian transplant centers. An annual report is finalized by combining these data with data from the Eurotransplant database. Results: An increase of both potential donors (n = 501, +14.4%) and effective donors (n = 273, +16.7%) was observed in 2006 versus 2005. Among effective donors, 28 were non-heart-beating donors (10.25%). Overall donor ratio was 26.26 donors per million inhabitants. Within potential donors, absence of organ harvesting was due to medical contraindications (28%), family refusal (13%), or legal refusal (2%). Donor mean age was 46.4 years and mean organs/donor was 3.21 ± 1.7. An overall reduction of Belgian waiting lists was observed in 2006 as compared with 2005 (-5.7% for kidney, -25.7% for liver, -9.4% for heart, -6.7% for lung, and -11.7% for pancreas), while waiting list mortality was 18% for liver, 11% for heart, and 7% for lung. As compared with 2005, transplant activities increased for kidney (n = 485, +24.3%), heart ± lungs (n = 73, +7.3%), and lungs (n = 83, +39.4%) but decreased for liver (n = 236, -2.1%). Living donation represented 8.45% for kidney (+28.1% vs 2005) and 8% for liver transplantation (-29.6%). Conclusion: Globally, a marked increase of procurement and transplant activities was observed in 2006, allowing to limit waiting list and waiting list mortality. Further increase of living donor activity and non-heart-beating donation remains necessary to extend the donor pool. © 2007 Elsevier Inc. All rights reserved.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

Boosting heterologous protein production in transgenic dicotyledonous seeds using Phaseolus vulgaris regulatory sequences

Over the past decade, several high-value proteins have been produced in different transgenic plant tissues such as leaves, tubers, and seeds(1,2). Despite recent advances, many heterologous proteins accumulate to low concentrations, and the optimization of expression cassettes to make in planta production and purification economically feasible remains critical. Here, the regulatory sequences of the seed storage protein gene arcelin 5-I (arc5-I) of common bean (Phaseolus vulgaris)(3) were evaluated for producing heterologous proteins in dicotyledonous seeds. The murine single-chain variable fragment (scFv) G4 (ref. 4) was chosen as model protein because of the current industrial interest in producing antibodies and derived fragments in crops(5,6). In transgenic Arabidopsis thaliana seed stocks, the scFv under control of the 35S promoter of the cauliflower mosaic virus (CaMV) accumulated to approximately 1% of total soluble protein (TSP). However, a set of seed storage promoter constructs boosted the scFv accumulation to exceptionally high concentrations, reaching no less than 36.5% of TSP in homozygous seeds. Even at these high concentrations, the scFv proteins had antigen-binding activity and affinity similar to those produced in Escherichia coli. The feasibility of heterologous protein production under control of arc5-I regulatory sequences was also demonstrated in Phaseolus acutifolius, a promising crop for large-scale production