Lutein and zeaxanthin are commonly referred to as the macular carotenoids, as they are localized to ocular tissues and their loss is associated with age-related macular degeneration. Thirty-four high-carotenoid (HiC) lines exhibiting uniquely high concentrations of carotenoids resulted from alíele mining of the Orange Flint race using traditional breeding techniques and visual selection for deep orange endosperm color. Total carotenoid concentrations of the HiC lines ranged from 50 to 101 pg g"^ dry weight (DW) with iutein and zeaxanthin concentrations as high as 66 and 65 pg g'' DW, respectively, levels higher than reported in previous germplasm surveys. The HiC lines fall into three classes based on the accumulation of the major carotenoid: "high-lutein," "high-zeaxanthin," and "balanced." Significant year effects were observed for carotenoid concentrations but not for profiles. During kernel development the pattern of carotenoid accumulation in the HiC lines did not appear to be different than in yellow corn belt dent lines. Collectively, the HiC lines represented only one yi haplotype, 10 unique haplotypes of IcyE, and two unique haplotypes at crtRBL Interestingly, previously identified diagnostic polymorphisms within IcyE did not appear to be useful in distinguishing between high-lutein and high-zeaxanthin HiC lines, and high .i-carotene levels were achieved despite the presence of a suboptimal crtRBi alíele. The HiC lines illustrate the utility of mining allelic variation from exotic sources coupled with the power of simple visual selection and the potential limitations of diagnostic polymorphisms.

Determination of the role of mitochondrial (mt) ATP synthesis in plant metabolism is complicated by chloroplastic ATP synthesis. To differentiate ATP synthesis from these two organelles, we created transgenic Arabidopsis plants in which two different subunits of the mt ATP synthase, the oligomycin sensitivity-conferring protein (OSCP) (=delta) (ATP5) and the gamma (ATP3) subunit, were expressed individually in antisense orientation under the control of a dexamethasone-inducible promoter. The phenotypic effects of antisense expression were identical for both atp5 and atp3. Seedling lethality resulted from induction during germination in the light, demonstrating the essentiality of both gene products. Reduced expression of either gene resulted in stunting of dark-grown (etiolated) seedlings, downward curling or wavy-edged leaf margins of light-grown plants and ball-shaped unexpanded flowers. Antisense induction reduced total ATP levels in dark-grown (etiolated) seedlings germinated on media lacking sucrose, but increased total ATP levels in induced light-grown plants and in induced dark-grown seedlings germinated on media containing sucrose. Induction reduced transcript levels for two transcription factors (TCP3 and TCP4) whose decreased expression is associated with a similar wavy-edged leaf phenotype in Arabidopsis, and increased transcript levels for dynamin-related proteins whose increased expression is associated with increased mt division. Reduced expression of these subunits of the mt ATP synthase is proposed to disturb cellular redox states, which ultimately manifest downstream as diverse and seemingly unrelated phenotypes.

A methodology is presented for the hydroponic cultivation of Arabidopsis thaliana (L.) Heynh. that is simple and inexpensive to construct from easily obtainable and reusable materials. High-quality and uniform plant material, free of algae and disease, are consistently produced through all life-stages. Minor modifications to previous protocols that reduce cost and labour include the use of small food-storage containers with snap-on lids, aluminum foil wrap to cover the containers to occlude light, commercial hydroponic fertilizer as nutrient solution, and clipped 1.5 mL microfuge tubes as plug holders. Seed is germinated on nutrient agar plates, instead of directly on rockwool plugs; after the seedlings have grown to the six-leaf stage, they are removed from the plates and the roots are enclosed within rockwool plugs that have been cut longitudinally. Subsequent seedling growth is very uniform and the elimination of oversowing and subsequent thinning results in a dramatic reduction in seed and seedling wastage.

Hydroponic cultivation systems provide convenient means of delivering chemical inducers of gene expression to transgenic plants. The glucocorticoid dexamethasone is a water-insoluble inducer and is usually prepared as a stock solution in an organic solvent before addition to the hydroponic nutrient solution. We investigated the effects of ethanol, methanol, and dimethyl sulfoxide (DMSO) on plant appearance and root bacterial growth in nonsterile hydroponic Arabidopsis thaliana (L.) Heynh. culture after 8 d of exposure to the solvent. Ethanol and methanol promoted root bacterial growth and visibly affected overall plant appearance at levels as low as 0.001%, and the effects increased in direct relation to alcohol concentration. DMSO promoted root bacterial growth to a lesser extent than the alcohols and plant appearance was negatively affected only at concentrations of DMSO above 0.01%, suggesting that DMSO may be the best solvent choice. We also demonstrated, however, that dexamethasone prepared as a suspension in water, without dissolution in an organic solvent, was completely effective at inducing transcription of the transgene, thus eliminating the potentially confounding effects of solvents in the interpretation of results.