A plant source of omega-3 fatty acid (FA) that can raise tissue eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA) is needed. A soybean oil (SBO) containing approximately 20% stearidonic acid [SDA; the delta-6 desaturase product of alpha-linolenic acid (ALA)] derived from genetically modified soybeans is under development. This study compared the effects of EPA to SDA-SBO on erythrocyte EPA+DHA levels (the omega-3 index). Overweight healthy volunteers (n=45) were randomized to SDA-SBO (24 ml/day providing approximately 3.7 g SDA) or to regular SBO (control group) without or with EPA ethyl esters (approximately 1 g/day) for 16 weeks. Serum lipids, blood pressure, heart rate, platelet function and safety laboratory tests were measured along with the omega-3 index. A per-protocol analysis was conducted on 33 subjects (11 per group). Compared to baseline, average omega-3 index levels increased 19.5% in the SDA group and 25.4% in the EPA group (p<0.05 for both, vs. control). DHA did not change in any group. Relative to EPA, SDA increased RBC EPA with about 17% efficiency. No other clinical endpoints were affected by SDA or EPA treatment (vs. control). In conclusion, SDA-enriched SBO significantly raised the omega-3 index. Since EPA supplementation has been shown to raise the omega-3 index and to lower risk for cardiac events, SDA-SBO may be a viable plant-based alternative for providing meaningful intakes of cardioprotective omega-3 FAs.

The safety of 5-enolpyruvylshikimate-3-phosphate synthase enzyme derived from Agrobacterium sp. strain CP4 (CP4 EPSPS) was assessed. CP4 EPSPS is the only protein introduced by genetic manipulation that is expressed in glyphosate-tolerant soybeans, which are being developed to provide new weed-control options for farmers. Expression of this protein in plants imparts high levels of glyphosate tolerance. The safety of CP4 EPSPS was ascertained by evaluating both physical and functional characteristics. CP4 EPSPS degrades readily in simulated gastric and intestinal fluids, suggesting that this protein will be degraded in the mammalian digestive tract upon ingestion as a component of food or feed, There were no deleterious effects due to the acute administration of CP4 EPSPS to mice by gavage at a high dosage of 572 mg/kg body wt, which exceeds 1000-fold tha anticipated consumption level of food products potentially containing CP4 EPSPS protein. CP4 EPSPS does not pose any important allergen concerns because this protein does not possess characteristics typical of allergenic proteins. These data, in combination with seed compositional analysis and animal feeding studies, support the conclusion that glyphosate-tolerant soybean are as safe and nutritious as traditional soybeans currently being marketed.

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Reports of abnormalities in vitamin D, calcium, and bone status associated with anticonvulsant use are inconsistent and difficult to interpret because of widely varying study designs, particularly for ambulatory status. We examined the relative effects of anticonvulsant use and ambulatory status on vitamin D, calcium, and bone status in a large group (n = 338) of children who had either normal motor function (ambulatory) or were nonambulatory and either receiving anticonvulsants or not; all had developmental delays. Data included diet records, serum analyses (calcium and calcidiol), and hand-wrist radiographs evaluated for bone maturation and quality. Data were analyzed by using a general linear models (GLM) procedure. Dietary and biochemical data were compared with those of a group of 34 normal children. There were no differences in calcium or vitamin D intakes among the four study groups; however, a high percentage of intakes was below the recommended dietary allowances for calcium (56%) and vitamin D (70%). Vitamin D intakes were positively associated with serum calcium (P < 0.005) and calcidiol (P < 0.01) concentrations. Analysis of covariance indicated that ambulatory status but neither anticonvulsant use nor their interaction contributed significantly to the prediction of serum calcium (P < 0.009) and calcidiol (P < 0.0001), the Z scores for number of ossified centers (P < 0.008), bone age (P < 0.0001), and bone area (P < 0.003). A strong interaction between anticonvulsant use and ambulatory status was seen for percentage cortical area (P < 0.0008), which was entirely due to anticonvulsant use in nonambulatory children (effect size = 0.98). Results suggest that ambulatory status is more important than was recognized previously in relation to abnormalities in vitamin D, calcium, and bone statuses; that all nonambulatory children may be at risk for low serum calcidiol and osteopenia; and that routine monitoring of risk and consideration of prophylactic vitamin D supplementation are warranted.

The lack of alternative selectable markers in crop transformation has been a substantial barrier for commercial application of agricultural biotechnology. We have developed an efficient selection system for wheat transformation using glyphosate-tolerant CP4 and GOX genes as a selectable marker. Immature embryos of the wheat cultivar Bobwhite were bombarded with two separate plasmids harboring the CP4/GOX and GUS genes. After a 1 week delay, the bombarded embryos were transferred to a selection medium containing 2 mM glyphosate. Embryo-derived calli were subcultured onto the same selection medium every 3 weeks consecutively for 9-12 weeks, and were then regenerated and rooted on selection media with lower glyphosate concentrations. Transgenic plants tolerant to glyphosate were recovered. ELISA assay confirmed expression of the CP4 and GOX genes in R0 plants. Southern blot analysis demonstrated that the transgenes were integrated into the wheat genomes and transmitted to the following generation. The use of CP4 and GOX genes as a selectable marker provides an efficient, effective, and alternative transformation selection system for wheat.

We tested two genes together in hybrid poplars (genus Populus), CP4 and GOX, for imparting tolerance to glyphosate (the active ingredient in Roundup® herbicide). Using Agrobacterium-based transformation, 80 independent transgenic lines (i.e., products of asexual gene transfer) were produced in a variety of hybrid poplar clones (40 lines in Populus trichocarpa Torr. & Gray × Populus deltoides Bartr., 35 lines in Populus tremula L. × Populus alba L., and five lines in P. tremula × Populus tremuloides Michx.). We evaluated glyphosate tolerance over 2 years in field studies conducted in eastern and western Oregon. Ten percent of our transgenic lines showed no foliar damage or reduced growth after being sprayed with Roundup® at concentrations above normal commercial rates. Lack of damage was associated with expression of the CP4 gene but not of the GOX gene. It was suspected that GOX caused undesirable side effects, so we produced 12 lines into which only the CP4 gene was inserted. The performance of these newly regenerated lines was compared with an identical number of lines, produced in the same genotype, that had previously been engineered to contain both CP4 and GOX. Growth of the lines transformed with just CP4 was significantly better than those containing both genes and exhibited less damage in response to glyphosate treatment. This is the first report of transgenic poplars exhibiting high levels of glyphosate tolerance when grown under field conditions. With a modest transformation effort, it is possible to produce lines with commercially useful levels of glyphosate tolerance and little apparent collateral genetic damage.

Genetically modified cotton lines have been developed that are tolerant to glyphosate, the active ingredient in the herbicide Roundup. The new lines were generated by Agrobacterium tumfaciens-mediated transfer of a gene encoding 5-enolpyruvylshikimate-3-phosphate synthase isolated from Agrobacterium sp. CP4 (CP4 EPSPS). Lines were screened via greenhouse spray tests and field evaluations to identify agronomically acceptable lines with a commercial level of tolerance to glyphosate. Two lines were characterized. Lines 1445 and 1698 were transformed with different vectors that encode for the CP4 EPSPS and the neomycin phosphotransferase II (NPTII) marker protein. Both lines contain a single DNA insertion that segregates in a typical Mendelian fashion. Line 1445 contains a single copy of the CP4 EPSPS gene, whereas the line 1698 contains two copies of the CP4 EPSPS gene at a single insertion site. The stability of each DNA insertion was demonstrated by Southern analysis across the R3 and R5 generations. The expression levels of the CP4 EPSPS and NPTII were quantitated by ELISA in leaf and seed samples collected in 1993 and 1994 field trials. The use of glyphosate-tolerant cotton will enable the grower to take advantage of additional weed management alternatives. Keywords: Cotton; genetically modified; herbicide tolerant; Roundup

It is widely recognized that the application of organic materials is one of the most effective ways of increasing soil organic carbon (SOC) levels and improving soil quality, but do all forms of organic matter input have the same impact on soil properties A network of seven experimental sites investigated the effects on soil quality of annual applications over a minimum of 3 years of compost and food-based digestate in comparison with farmyard manure (FYM) and livestock slurry. Two of the sites were existing experimental platforms which had previously benefitted from applications of FYM, livestock slurry and green compost allowing the effects of longer-term applications (6-17 years) on soil properties to be quantified. The application of all organic materials increased soil nutrient supply (total nitrogen, extractable phosphorus, potassium, and magnesium) within a short timescale (<3 years), whereas SOC contents were only increased following the long-term (9 years or more) application of bulky organic materials (compost and FYM). SOC increases were associated with improvements in soil biological (microbial biomass) and physical properties (reduced bulk density), although the level of improvement was dependent on the quality of the organic material applied (as determined by its lignin content, an indicator of resistance to decomposition). Applications of low dry matter content materials (digestates and livestock slurries) had a limited capacity to improve soil biological and physical functioning, due to their low organic matter loading.