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Screening tests in which 30 species of succulent plants were inoculated with A136 or C58 strains of Agrobacterium tumefaciens led to selecting genus Kalanchoe for further study of Agrobacterium-mediated transformation. Regeneration studies showed that 4.55 µM Thidiazuron (TDZ) combined with 2.15 µM α-naphtaleneacetic acid (NAA) significantly proliferated callus development from Kalanchoe pinnata leaf explants in Murashige and Skoog (MS) medium. Subsequently, 4.55 µM TDZ significantly enhanced shoot formation from the green and compact callus. NAA (5.37 µM) accelerated root growth leading to establishment of transgenic plants in soil. A gene encoding the fluorescent protein, ZsGreen1, was cloned into a plasmid vector harboring a constitutive figwort mosaic virus (FMV) 35S promoter, a kanamycin resistance gene, and a NOS-terminator. The plasmids were delivered into the genome of K. pinnata by A. tumefaciens strain C58 (ABI). The ZsGreen1-transgenic plants exhibited very strong, green fluorescence from the callus, leaves, stems, roots, and flowers visible through a green emission filter (GFP, 480/510 nm, excitation/emission) under excitation by ultra-violet light. Transcription analysis by sqRT-PCR and qPCR confirmed the stable integration of the ZsGreen1 in the genome of K. pinnata. The FMV 35S promoter conferred uniform expression throughout whole plant organ. ZsGreen1 was proven to be a stable marker for gene expression in genetic transformation of K. pinnata. The ZsGreen1-transgenic Kalanchoe pinnata mediated by Agrobacterium tumefaciens exhibited strong, green fluorescence from the callus, leaves, stems, roots and flowers under the control of the figwort mosaic virus (FMV) 35S promoter.

BackgroundRNA sequencing has been widely used to profile genome-wide gene expression and identify candidate genes controlling disease resistance and other important traits in plants. Gerbera daisy is one of the most important flowers in the global floricultural trade, and powdery mildew (PM) is the most important disease of gerbera. Genetic improvement of gerbera PM resistance has become a crucial goal in gerbera breeding. A better understanding of the genetic control of gerbera resistance to PM can expedite the development of PM-resistant cultivars.ResultsThe objectives of this study were to identify gerbera genotypes with contrasting phenotypes in PM resistance and sequence and analyze their leaf transcriptomes to identify disease resistance and susceptibility genes differentially expressed and associated with PM resistance. An additional objective was to identify SNPs and SSRs for use in future genetic studies. We identified two gerbera genotypes, UFGE 4033 and 06–245-03, that were resistant and susceptible to PM, respectively. De novo assembly of their leaf transcriptomes using four complementary pipelines resulted in 145,348 transcripts with a N50 of 1124 bp, of which 67,312 transcripts contained open reading frames and 48,268 were expressed in both genotypes. A total of 494 transcripts were likely involved in disease resistance, and 17 and 24 transcripts were up- and down-regulated, respectively, in UFGE 4033 compared to 06–245-03. These gerbera disease resistance transcripts were most similar to the NBS-LRR class of plant resistance genes conferring resistance to various pathogens in plants. Four disease susceptibility transcripts (MLO-like) were expressed only or highly expressed in 06–245-03, offering excellent candidate targets for gene editing for PM resistance in gerbera. A total of 449,897 SNPs and 19,393 SSRs were revealed in the gerbera transcriptomes, which can be a valuable resource for developing new molecular markers.ConclusionThis study represents the first transcriptomic analysis of gerbera PM resistance, a highly important yet complex trait in a globally important floral crop. The differentially expressed disease resistance and susceptibility transcripts identified provide excellent targets for development of molecular markers and genetic maps, cloning of disease resistance genes, or targeted mutagenesis of disease susceptibility genes for PM resistance in gerbera.

Fluorescent proteins (FT) have become essential, biological research tools. Many novel genes have been cloned from a variety of species and modified for effective, stable, and strong expression in transgenic organisms. Although there are many applications, FT expression has been employed most commonly at the cellular level in plants. To investigate FT expression at the whole-plant level, particularly in flowers, petunia ‘Mitchell Diploid’ [MD (Petunia ×hybrida)] was genetically transformed with seven genes encoding FTs: DsRed2, E2Crimson, TurboRFP, ZsGreen1, ZsYellow1, rpulFKz1, or aeCP597. Each gene was cloned into a pHK-DEST-OE vector harboring constitutive figwort mosaic virus 35S promoter and NOS-terminator. These plasmids were individually introduced into the genome of MD by Agrobacterium tumefaciens–mediated transformation. Shoot regeneration efficiency from the cocultured explants ranged from 8.3% to 20.3%. Various intensities of red, green, and yellow fluorescence were detected from TurboRFP, ZsGreen1, and ZsYellow1-transgenic flowers, respectively, under ultraviolet light for specific excitation and emission filters. More than 70% of plants established from the regenerated shoots were confirmed as transgenic plants. Transgenic ZsGreen1 petunia generated strong, green fluorescence in all flower organs of T0 plants including petals, stigmas, styles, anthers, and filaments. Most of the chromophores were localized to the cytoplasm but also went into the nuclei of petal cells. There was a positive linear relationship (R2 = 0.88) between the transgene expression levels and the relative fluorescent intensities of the ZsGreen1-transgenic flowers. No fluorescence was detected from the flowers of DsRed2-, E2Crimson-, rpulFKz1-, or aeCP597-transgenic petunias even though their gene transcripts were confirmed through semiquantitative reverse transcriptase-polymerase chain reaction. T1 generation ZsGreen1 plants showed green fluorescence emission from the cotyledons, hypocotyls, and radicles, which indicated stable FT expression was heritable. Four homozygous T2 inbred lines were finally selected. Throughout this study, we demonstrated that ZsGreen1 was most suitable for generating visible fluorescence in MD flowers among the seven genes tested. Thus, ZsGreen1 may have excellent potential for better utility as a sensitive selectable marker.

Petunia × hybrida cv ‘Mitchell Diploid’ floral volatile benzenoid/phenylpropanoid (FVBP) biosynthesis ultimately produces floral volatiles derived sequentially from phenylalanine, cinnamic acid, and p-coumaric acid. In an attempt to better understand biochemical steps after p-coumaric acid production, we cloned and characterized three petunia transcripts with high similarity to p-coumarate 3-hydroxylase (C3H), hydroxycinnamoyl-CoA:shikimate/quinate hydroxycinnamoyl transferase (HCT), and caffeoyl shikimate esterase (CSE). Transcript accumulation of PhC3H and PhHCT was highest in flower limb tissue during open flower stages. PhCSE transcript accumulation was also highest in flower limb tissue, but it was detected earlier at initial flower opening with a bell-shaped distribution pattern. Down regulation of endogenous PhC3H transcript resulted in altered transcript accumulation of many other FVBP network transcripts, a reduction in floral volatiles, and the emission of a novel floral volatile. Down regulation of PhHCT transcript did not have as large of an effect on floral volatiles as was observed for PhC3H down regulation, but eugenol and isoeugenol emissions were significantly reduced on the downstream floral volatiles. Together these results indicate that PhC3H is involved in FVBP biosynthesis and the reduction of PhC3H transcript influences FVBP metabolism at the network level. Additional research is required to illustrate PhHCT and PhCSE functions of petunia.

Coleus (Plectranthus scutellarioides) is an attractive and popular ornamental plant with propagation mainly achieved through vegetative cuttings. For commercial purposes, it is of interest to enhance the speed of establishment while maintaining high quality. Light quality has been shown to influence adventitious root development, so these experiments examined the effect of narrow-bandwidth light treatments on root growth and overall plant quality for seven coleus cultivars with vegetative cuttings in potting soil and one cultivar with shoot tip in vitro cultures onto Murashige and Skoog (MS) agar medium. During the 28 days of the propagation period, the cuttings grown under narrow-bandwidth red light (R; 663.4 nm at peak) more than doubled in the adventitious root number compared with those under blue light (B; 445.7 nm at peak) and green light (G; 530.0 nm at peak) in five cultivars. R light also increased fresh weight of the cuttings by 55.6% more than G light. In comparison, the cuttings grown under G light yielded significantly lower root and shoot dry mass than other light treatments. R light cuttings showed more dry mass content (9.63%) than those under white light (W; 437.4 nm and 559.5 nm at peak) and G light (7.85% and 5.86%, respectively). A positive correlation (R2 = 0.598, P < 0.001) was found between the formation of adventitious roots and gained fresh weight of cuttings. R light made the reddish color of leaves significantly stronger in most cultivars, whereas the cuttings exposed to G light became less vivid compared with other light conditions. When the shoot tips were propagated in vitro onto MS medium, R light treatment initiated the root development more rapidly than other lights, with significantly greater rooting rate (20.0% and 63.6%, respectively) at day 5 and 10. The shoot tips under R light also formed significantly more roots (12.3 per cutting) than those grown under narrow-bandwidth B light (5.8 per cutting). The shoot tips showed browning at an early stage and newly emerged leaves grew very compactly under B light. The combination of red and green light (R+G) increased more than twice as much roots and dry mass compared with W light. In addition, the R+G light led to morphological changes, including larger leaves and longer petioles and internodes than those in other light treatments. The exposure to R+G+B and B light made the shoots very compact for the 28 days of in vitro culture period and significantly increased the chlorophyll contents resulting in dark green leaves.

Seed germination and seedling vigor can be affected by environmental cues experienced by the mother plant. However, information about how the maternal environment affects seed quality is scarce in ornamental plants. This study aimed to investigate the effects of two different maternal environments on the seed germination and seedling vigor of Petunia × hybrida under a variety of abiotic stresses. Petunia mother plants were grown in either a greenhouse during the summer months or an indoor controlled-temperature-and-light environment. Collected seeds were subjected to external stressors, including polyethylene glycol (PEG), sodium chloride (NaCl), high temperature, and abscisic acid (ABA), to determine seed germination percentage and seedling vigor. Results indicated that seeds harvested from the mother plants grown in a controlled environment germinated better than seeds harvested from the mother plants grown in the greenhouse when suboptimal germination conditions were applied. Additionally, the seedlings from the controlled maternal environment performed better in both ABA and salinity stress tests than the greenhouse seedlings. Interestingly, the greenhouse seedlings displayed less reactive oxygen species (ROS) damage and lower electrolyte leakage than the controlled environment seedlings under dehydration stress. The difference in germination and seedling vigor of seeds from the two different maternal environments might be due to the epigenetic memory inherited from the mother plants. This study highlighted the strong impact of the maternal environment on seed germination and seedling vigor in Petunia and may assist in high-quality seed production in ornamental plants.

SummaryColeus is a popular ornamental plant that exhibits a diverse array of foliar color patterns. New cultivars are currently hand selected by both amateur and experienced plant breeders. In this study, we reimagine coleus breeding using a quantitative color analysis framework.Despite impressive advances in high-throughput data collection and processing, complex color patterns remain challenging to extract from image datasets. Using a new phenotyping approach called “ColourQuant,” we extract and analyze pigmentation patterns from one of the largest coleus breeding populations in the world.Working with this massive dataset, we are able to analyze quantitative relationships between maternal plants and their progeny, identify features that underlie breeder-selections, and collect and compare consumer input on trait preferences.This study is one of the most comprehensive explorations into complex color patterning in plant biology and provides new insights and tools for exploring the color pallet of the plant kingdom.

Polyphenol oxidase (PPO) is a well-characterized enzyme that is conserved across plant species and is responsible for catalyzing the rapid polymerization of o-quinones that results in tissue browning. As these oxidation reactions result in significant reductions in food crop commercial viability, nutritional content, and general desirability, an opportunity exists to improve a variety of crops via genetic regulation of PPO expression. The main objective of this project was to clone the DNA sequence of genes encoding PPO. Using mRNA as a template to synthesize DNA, two putative PPO sequences were cloned from Ocimum basilicum and Ocimum americanum, two basil species of commercial relevance. A secondary objective of this project was to insert a cloned PPO sequence into an RNAi vector for the purpose of making transgenic basil plants with reduced PPO expression. A hairpin RNAi construct was successfully developed to reduce the expression of both PPO genes as their DNA sequences were highly homologous. The next step for this project will depend on the development of transformation and regeneration protocols for these species, which have proven to be difficult. Once the RNAi PPO constructs are introduced into basil cells and successfully regenerated into plants, reduced browning of basil leaf tissue when it is wounded or cut is expected.

Accurate diagnosis of mild cognitive impairment (MCI) before conversion to Alzheimer’s disease (AD) is invaluable for patient treatment. Many works showed that MCI and AD affect functional and structural connections between brain regions as well as the shape of cortical regions. However, ‘shape connections’ between brain regions are rarely investigated -e.g., how morphological attributes such as cortical thickness and sulcal depth of a specific brain region change in relation to morphological attributes in other regions. To fill this gap, we unprecedentedly design morphological brain multiplexes for late MCI/AD classification. Specifically, we use structural T1-w MRI to define morphological brain networks, each quantifying similarity in morphology between different cortical regions for a specific cortical attribute. Then, we define a brain multiplex where each intra-layer represents the morphological connectivity network of a specific cortical attribute, and each inter-layer encodes the similarity between two consecutive intra-layers. A significant performance gain is achieved when using the multiplex architecture in comparison to other conventional network analysis architectures. We also leverage this architecture to discover morphological connectional biomarkers fingerprinting the difference between late MCI and AD stages, which included the right entorhinal cortex and right caudal middle frontal gyrus.

International audience; We identified rare coding variants associated with Alzheimer's disease in a three-stage case-control study of 85,133 subjects. In stage 1, we genotyped 34,174 samples using a whole-exome microarray. In stage 2, we tested associated variants (P < 1 × 10-4) in 35,962 independent samples using de novo genotyping and imputed genotypes. In stage 3, we used an additional 14,997 samples to test the most significant stage 2 associations (P < 5 × 10-8) using imputed genotypes. We observed three new genome-wide significant nonsynonymous variants associated with Alzheimer's disease: a protective variant in PLCG2 (rs72824905: p.Pro522Arg, P = 5.38 × 10-10, odds ratio (OR) = 0.68, minor allele frequency (MAF)cases = 0.0059, MAFcontrols = 0.0093), a risk variant in ABI3 (rs616338: p.Ser209Phe, P = 4.56 × 10-10, OR = 1.43, MAFcases = 0.011, MAFcontrols = 0.008), and a new genome-wide significant variant in TREM2 (rs143332484: p.Arg62His, P = 1.55 × 10-14, OR = 1.67, MAFcases = 0.0143, MAFcontrols = 0.0089), a known susceptibility gene for Alzheimer's disease. These protein-altering changes are in genes highly expressed in microglia and highlight an immune-related protein-protein interaction network enriched for previously identified risk genes in Alzheimer's disease. These genetic findings provide additional evidence that the microglia-mediated innate immune response contributes directly to the development of Alzheimer's disease.

Brain imaging genetics intends to uncover associations between genetic markers and neuroimaging quantitative traits. Sparse canonical correlation analysis (SCCA) can discover bi-multivariate associations and select relevant features, and is becoming popular in imaging genetic studies. The L1-norm function is not only convex, but also singular at the origin, which is a necessary condition for sparsity. Thus most SCCA methods impose $${\ell }_{{\bf{1}}}$$ ℓ 1 -norm onto the individual feature or the structure level of features to pursuit corresponding sparsity. However, the $${\ell }_{{\bf{1}}}$$ ℓ 1 -norm penalty over-penalizes large coefficients and may incurs estimation bias. A number of non-convex penalties are proposed to reduce the estimation bias in regression tasks. But using them in SCCA remains largely unexplored. In this paper, we design a unified non-convex SCCA model, based on seven non-convex functions, for unbiased estimation and stable feature selection simultaneously. We also propose an efficient optimization algorithm. The proposed method obtains both higher correlation coefficients and better canonical loading patterns. Specifically, these SCCA methods with non-convex penalties discover a strong association between the APOE e4 rs429358 SNP and the hippocampus region of the brain. They both are Alzheimer’s disease related biomarkers, indicating the potential and power of the non-convex methods in brain imaging genetics.

Lantana camara L., a popular nursery and landscape plant, is categorized as an invasive species in Florida, because it produces viable pollen and cross-pollinates with the native species Lantana depressa Small. The invasive potential of L. camara is a challenging issue for the nursery and landscape industry, so sterile non-invasive cultivars are needed to replace fertile invasive ones. This study aimed to determine the ploidy level and male fertility of both commercial L. camara cultivars and breeding lines to identify male-sterile cultivars and assess the effectiveness of sterile triploid production in L. camara. A polyploid series was identified among 32 L. camara cultivars and breeding lines. Male fertility, based on pollen stainability, varied widely among the cultivars/breeding lines. Ploidy level was the most important factor determining L. camara pollen stainability/male sterility. On average, diploids exhibited the highest pollen stainability (64.6%) followed by tetraploids (45.1%), pentaploids (34.6%), and hexaploids (18.0%). Triploids showed the lowest pollen stainability (9.3%), suggesting that generating triploids would be an effective genetic approach to producing sterile L. camara and reducing its pollen-mediated invasiveness. Pollen stainability of triploid cultivars, Balandpawn (LandmarkTM Pink Dawn PP15,516), Lemon Drop, Miss Huff, New Gold, New Red Lantana, Red Butler, Red Spread Lantana, Samson Lantana, and Sunset Lantana was consistently below 10%. A number of triploid cultivars had pollen stainability approaching 20% to 30%, indicating a necessity for careful examination and screening of newly produced triploids to ensure high sterility in selected triploids. Pollen stainability variation was observed within ploidy levels, implying the existence of other genetic and environmental factors that influence the pollen stainability/male fertility of L. camara. Results from this study suggest that there is excellent potential to develop genetically sterile cultivars in L. camara for the U.S. nursery and landscape industry.

Floral volatile benzenoid/phenylpropanoid (FVBP) biosynthesis consists of numerous enzymatic and regulatory processes. The initial enzymatic step bridging primary metabolism to secondary metabolism is the condensation of phosphoenolpyruvate (PEP) and erythrose-4-phosphate (E4P) carried out via 3-DEOXY-D-ARABINO-HEPTULOSONATE-7-PHOSPHATE (DAHP) synthase. Here, identified, cloned, localized, and functionally characterized were two DAHP synthases from the model plant species Petunia × hybrida cv ‘Mitchell Diploid’ (MD). Full-length transcript sequences for PhDAHP1 and PhDAHP2 were identified and cloned using cDNA SMART libraries constructed from pooled MD corolla and leaf total RNA. Predicted amino acid sequence of PhDAHP1 and PhDAHP2 proteins were 76% and 80% identical to AtDAHP1 and AtDAHP2 from Arabidopsis, respectively. PhDAHP1 transcript accumulated to relatively highest levels in petal limb and tube tissues, while PhDAHP2 accumulated to highest levels in leaf and stem tissues. Through floral development, PhDAHP1 transcript accumulated to highest levels during open flower stages, and PhDAHP2 transcript remained constitutive throughout. Radiolabeled PhDAHP1 and PhDAHP2 proteins localized to plastids, however, PhDAHP2 localization appeared less efficient. PhDAHP1 RNAi knockdown petunia lines were reduced in total FVBP emission compared to MD, while PhDAHP2 RNAi lines emitted ‘wildtype’ FVBP levels. These results demonstrate that PhDAHP1 is the principal DAHP synthase protein responsible for the coupling of metabolites from primary metabolism to secondary metabolism, and the ultimate biosynthesis of FVBPs in the MD flower.

Light intensity, duration, direction, and wavelength are informative to plants. The biochemical circuits that connect specific light wavelengths to expression of specific genes and the metabolic networks they govern have been well defined. However, little emphasis has been placed on how discrete wavelengths of light, alone or in combination, may be applied to manipulate postharvest qualities of high-value horticultural crops. Using narrow-bandwidth LED light we test the hypothesis that discrete light wavelengths can affect the accumulation of volatile compounds known to affect aroma or taste in select flower and fruit products. Volatile benzenoid/phenylpropanoid emission from petunia flowers could be altered with light application. Levels of a key floral volatile, 2-phenylethanol, increased with a red and far-red light treatment. Similar experiments demonstrated that fruit volatile profiles of tomato, strawberry, and blueberry can be manipulated with specific light treatments. These results suggest that compounds affecting sensory qualities of flowers and fruits can be modified by adjustment of ambient light conditions. These findings open new areas of inquiry about how the fragrance and flavor of flowers and fruits may be improved with simple changes in postharvest light conditions.

Previously, when selecting for flavor in the University of Florida southern highbush blueberry (SHB, Vaccinium corymbosum L. hybrids) breeding program, sugar/acid ratios and breeder preference were the only factors considered. A more precise method of evaluating flavor would include volatile compounds that may also contribute to the flavor experience. Therefore, volatile profiles of five SHB cultivars (Farthing, FL01-173, Scintilla, Star, and Sweetcrisp) were compared using gas chromatography–mass spectrometry. All cultivars were harvested on four separate dates within the harvest season, and fruit from each cultivar were also harvested at four developmental stages on the first harvest date. Among the cultivars, soluble solids content and volatile production tended to increase with fruit maturity, whereas titratable acidity decreased. All volatile components were more variable than measures of sugars and acids during the harvest season. Many of the volatiles present varied significantly between harvest dates, resulting in significant genotype × environment interactions during the harvest season. A closer examination of linalool, trans-2-hexenol, trans-2-hexenal, hexanal, and 1-penten-3-ol, five volatile compounds commonly associated with blueberry flavor, showed cultivar, developmental stage, and harvest date differences for each volatile. ‘Star’ experienced the least variation through the harvest period.

Flowers recruit floral visitors for pollination services by emitting fragrances. These scent signals can be intercepted by antagonists such as florivores to locate host plants. Hence, as a consequence of interactions with both mutualists and antagonists, floral bouquets likely consist of both attractive and defensive components. While the attractive functions of floral bouquets have been studied, their defensive function has not, and field-based evidence for the deterrence of floral-scent constituents is lacking. In field and glasshouse experiments with five lines of transgenic Petunia x hybrida plants specifically silenced in their ability to release particular components of their floral volatile bouquet, we demonstrate that the emission of single floral-scent compounds can dramatically decrease damage from generalist florivores. While some compounds are used in host location, others prevent florivory. We conclude that the complex blends that comprise floral scents are likely sculpted by the selective pressures of both pollinators and herbivores.

Pancreatitis is a complex, progressively destructive inflammatory disorder. Alcohol was long thought to be the primary causative agent, but genetic contributions have been of interest since the discovery that rare PRSS1, CFTR and SPINK1 variants were associated with pancreatitis risk. We now report two associations at genome-wide significance identified and replicated at PRSS1-PRSS2 (P < 1 × 10(-12)) and X-linked CLDN2 (P < 1 × 10(-21)) through a two-stage genome-wide study (stage 1: 676 cases and 4,507 controls; stage 2: 910 cases and 4,170 controls). The PRSS1 variant likely affects disease susceptibility by altering expression of the primary trypsinogen gene. The CLDN2 risk allele is associated with atypical localization of claudin-2 in pancreatic acinar cells. The homozygous (or hemizygous in males) CLDN2 genotype confers the greatest risk, and its alleles interact with alcohol consumption to amplify risk. These results could partially explain the high frequency of alcohol-related pancreatitis in men (male hemizygote frequency is 0.26, whereas female homozygote frequency is 0.07).

Summary Although human perception of food flavors involves integration of multiple sensory inputs, the most salient sensations are taste and olfaction [1]. Ortho- and retronasal olfaction are particularly crucial to flavor because they provide the qualitative diversity so important to identify safe versus dangerous foods [2]. Historically, flavor research has prioritized aroma volatiles present at levels exceeding the orthonasally measured odor threshold [3], ignoring the variation in the rate at which odor intensities grow above threshold. Furthermore, the chemical composition of a food in itself tells us very little about whether or not that food will be liked. Clearly, alternative approaches are needed to elucidate flavor chemistry. Here we use targeted metabolomics and natural variation in flavor-associated sugars, acids, and aroma volatiles to evaluate the chemistry of tomato fruits, creating a predictive and testable model of liking. This nontraditional approach provides novel insights into flavor chemistry, the interactions between taste and retronasal olfaction, and a paradigm for enhancing liking of natural products. Some of the most abundant volatiles do not contribute to consumer liking, whereas other less abundant ones do. Aroma volatiles make contributions to perceived sweetness independent of sugar concentration, suggesting a novel way to increase perception of sweetness without adding sugar.

Many elderly individuals remain dementia-free throughout their life. However, some of these individuals exhibit Alzheimer disease neuropathology on autopsy, evidenced by neurofibrillary tangles (NFTs) in AD-specific brain regions. We conducted a genome-wide association study to identify genetic mechanisms that distinguish non-demented elderly with a heavy NFT burden from those with a low NFT burden. The study included 344 non-demented subjects with autopsy (201 subjects with low and 143 with high NFT levels). Both a genotype test, using logistic regression, and an allele test provided genome-wide significant evidence that variants in the RELNgene are associated with neuropathology in the context of cognitive health. Immunohistochemical data for reelin expression in AD-related brain regions added support for these findings. Reelin signaling pathways modulate phosphorylation of tau, the major component of NFTs, either directly or through β-amyloid pathways that influence tau phosphorylation. Our findings suggest that up-regulation of reelin may be a compensatory response to tau-related or beta-amyloid stress associated with AD even prior to the onset of dementia.

R2R3-MYB transcription factors (TFs) are involved in diverse aspects of plant biology. Recently an R2R3-MYB was identified in Petunia x hybrida line P720 to have a role in the transcriptional regulation of floral volatile production. We propose a more foundational role for the R2R3-MYB TF EMISSION OF BENZENOIDS II (EOBII). The homolog of EOBII was isolated and characterized from P. x hybrida ‘Mitchell Diploid’ (MD) and Nicotiana attenuata. For both MD and N. attenuata, EOBII transcript accumulates to high levels in floral tissue with maximum accumulation at flower opening. When EOBII transcript levels are severely reduced using a stable RNAi (ir) approach in MD and N. attenuata, ir-EOBII flowers fail to enter anthesis and prematurely senesce. Transcript accumulation analysis demonstrated core phenylpropanoid pathway transcripts and cell wall modifier transcript levels are altered in ir-EOBII flowers. These flowers can be partially complemented by feeding with a sucrose, t-cinnamic acid, and gibberellic acid solution; presumably restoring cellular aspects sufficient for flower opening. Additionally, if ethylene sensitivity is blocked in either MD or N. attenuata, ir-EOBII flowers enter anthesis. These experiments demonstrate one R2R3-MYB TF can control a highly dynamic process fundamental to sexual reproduction in angiosperms: the opening of flowers.

Pantothenate kinase-associated neurodegeneration is a form of neurodegeneration with brain iron accumulation, characterized by a progressive movement disorder and prominent iron deposition in the globus pallidus. Formerly referred to as Hallervorden–Spatz syndrome, the disorder was renamed pantothenate kinase-associated neurodegeneration after discovery of the causative gene, PANK2. Although the pathological features of clinically characterized Hallervorden–Spatz syndrome have been described, the literature is confounded by the historical use of this term for nearly all conditions with prominent basal ganglia iron accumulation and by the fact that this term encompasses a genetically heterogeneous group of disorders, now referred to as ‘neurodegeneration with brain iron accumulation’. As a result, interpreting reports that precede molecular characterization of specific forms of neurodegeneration with brain iron accumulation is problematic. In the present studies, we describe neuropathological findings in six cases of molecularly confirmed pantothenate kinase-associated neurodegeneration. We identify prominent ubiquinated deposits in pantothenate kinase-associated neurodegeneration. We also characterize two distinct origins of spheroid bodies and delineate histological features of iron deposition. In so doing, we characterize fundamental features of the disease and redefine its nosological relationship to other neurodegenerative disorders.

The Alzheimer Disease Genetics Consortium (ADGC) performed a genome-wide association study of late-onset Alzheimer disease using a three-stage design consisting of a discovery stage (stage 1) and two replication stages (stages 2 and 3). Both joint analysis and meta-analysis approaches were used. We obtained genome-wide significant results at MS4A4A (rs4938933; stages 1 and 2, meta-analysis P (P(M)) = 1.7 × 10(-9), joint analysis P (P(J)) = 1.7 × 10(-9); stages 1, 2 and 3, P(M) = 8.2 × 10(-12)), CD2AP (rs9349407; stages 1, 2 and 3, P(M) = 8.6 × 10(-9)), EPHA1 (rs11767557; stages 1, 2 and 3, P(M) = 6.0 × 10(-10)) and CD33 (rs3865444; stages 1, 2 and 3, P(M) = 1.6 × 10(-9)). We also replicated previous associations at CR1 (rs6701713; P(M) = 4.6 × 10(-10), P(J) = 5.2 × 10(-11)), CLU (rs1532278; P(M) = 8.3 × 10(-8), P(J) = 1.9 × 10(-8)), BIN1 (rs7561528; P(M) = 4.0 × 10(-14), P(J) = 5.2 × 10(-14)) and PICALM (rs561655; P(M) = 7.0 × 10(-11), P(J) = 1.0 × 10(-10)), but not at EXOC3L2, to late-onset Alzheimer's disease susceptibility.

Floral volatiles are biologically and economically important plant derived chemical compounds. In petunia, floral volatile benzenoid/phenylpropanoid (FVBP) biosynthesis is controlled spatially, developmentally, and hormonally at molecular, metabolic, and biochemical levels. Over the last years, numerous genes have been shown to encode proteins that either directly catalyze a biochemical reaction yielding FVBP compounds, or are involved in metabolite flux prior to the formation of FVBP compounds. This FVBP gene network is specifically and coordinately transcribed. Multiple R2R3-MYB transcription factors are involved in the regulation of genes in the core metabolic pathways leading to a very unique mixture of emitted floral volatiles. The molecular puzzle is not complete, since the functions of the few FVBP transcription factors identified to date do not fully explain the transcriptional regulation of the entire gene network.

The relationship between fragrance and vase life and the role of ethylene on volatile emission in cut rose flowers was investigated. No relationship was observed between the amounts of volatile compounds emitted and vase life when fragrant and non-fragrant rose cultivars were compared. Neither ethylene production nor respiration rate of flowers was directly related with vase life. Volatile production during vase life was differential and independent among volatiles originating from different biosynthetic groups. Ethylene did not play a role in the regulation of volatile emission in rose flowers. Endogenous ethylene production was very low in most of the cultivars and did not show autocatalytic production trends. Volatile emission patterns during vase life did not parallel endogenous ethylene production. Exogenous ethylene exposure had differential effects among all cultivars, regardless of the fragrance of the flower. Fragrant cultivar ‘Osiana’ was highly sensitive to exogenous ethylene, with petals abscising within 24 h of ethylene (1 LL −1) exposure while other fragrant cultivars ‘Erin’ and ‘Lovely Dream’ had low ethylene sensitivity. Volatile production was unaffected by exogenous ethylene. The results of this study indicate that volatile emission in cut roses is not regulated by endogenous or exogenous ethylene and occurs independently of petal senescence and/or abscission. These results provide a better understanding of the complexity of volatile emission in rose flowers. © 2010 Elsevier B.V. All rights reserved.

BACKGROUND: Tomatoes contain high levels of several carotenoids including lycopene and β-carotene. Beyond their functions as colorants and nutrients, carotenoids are precursors for important volatile flavor compounds. In order to assess the importance of apocarotenoid volatiles in flavor perception and acceptability, we conducted sensory evaluations of near-isogenic carotenoid biosynthetic mutants and their parent, Ailsa Craig. RESULTS: The carotenoid contents of these tomatoes were extremely low in the r mutant, increased in lycopene in old gold, and higher in tetra-cis-lycopene and ζ-carotene in tangerine. The volatiles derived from these carotenoids (β-ionone, geranylacetone and 6-methyl-5-hepten-2-one) were proportionally altered relative to their precursors. Fruits were also analyzed for soluble solids, sugars, acids and flavor volatiles. Consumer panels rated the r mutant lowest for all sensory attributes, while Ailsa Craig was generally rated highest. Old gold and tangerine were rated intermediate in two of the three harvests. CONCLUSIONS: Several chemicals were negatively correlated with at least one of the hedonic scores while several others were positively correlated with tomato flavor acceptability. The results permitted identification of positive and negative interactions of volatiles with tomato flavor. Copyright © 2010 Society of Chemical Industry

Petunia (Petunia x hybrida cv 'Mitchell Diploid' [MD]) flowers emit high levels of multiple floral volatile benzenoid/phenylpropanoid (FVBP) compounds from anthesis to senescence in a concerted manner. Here we show seven genes responsible for the production of emitted FVBPs share similar transcript accumulation profiles through an analysis of four expression criteria. As a group, the FVBP gene transcripts accumulate to high levels in petal limb tissue of MD flowers from anthesis to senescence. Two to four hours of exogenous ethylene exposure reduces transcript levels of all FVBP genes examined, but 2h of treatment will not accelerate senescence or reduce volatile emissions in MD flowers. The FVBP genes show two obvious rhythmic patterns of transcript accumulation; however, corresponding enzyme activities of a subset of FVBP gene products do not. Together, these results depict floral volatile benzenoid/phenylpropanoid biosynthesis as a specific system with multiple regulatory features. One such feature is the highly regulated transcript accumulation of the FVBP genes. Additionally, ethylene may have a regulatory role in the FVBP system prior to a floral senescence program.

Ethylene is a gaseous plant hormone involved in many physiological processes including senescence, fruit ripening, and defence. Here the effects of pollination and wound-induced ethylene signals on transcript accumulation of benzoyl CoA:benzyl alcohol/phenylethanol benzoyltransferase (PhBPBT) are shown in Petuniaxhybrida cv. Mitchell 'Diploid' (MD). In petunia, PhBPBT is responsible for the biosynthesis of both benzyl benzoate and phenylethyl benzoate from benzyl alcohol and phenylethanol, respectively. RNAi-silenced lines, with reduced PhBPBT transcript, displayed reduced benzyl benzoate emission, and increased benzyl alcohol levels. Detailed expression analysis showed that PhBPBT is regulated by both light and an endogenous circadian rhythm, while it is also differentially regulated in response to ethylene in a tissue-specific manner. Twenty-four hours following pollination of MD flowers, expression of PhBPBT decreases in the corolla, while it increases in the ovary after 48 h. This is caused by ethylene that is emitted from the flower coinciding with fertilization as this is not observed in transgenic ethylene-insensitive plants (CaMV35S::etr1-1; 44568). Ethylene is also emitted from vegetative tissue of petunia following mechanical wounding, resulting in an increase in PhBPBT expression in the leaves where expression is normally below detection levels. Indicative of this pattern of expression, we hypothesize that PhBPBT and subsequent benzyl benzoate production is involved in defence-related processes in the corolla prior to pollination, in the ovary immediately following fertilization, and in vegetative tissue in response to wounding.

Hybrid Lilium (common name lily) cultivars are among the top produced domestic fresh cut flowers and potted plants in the US today. Many hybrid Lilium cultivars produce large and showy flowers that emit copious amounts of volatile molecules, which can negatively affect a consumer's appreciation or limit use of the plant product. There are few publications focused on the biochemistry, genetics, and/or molecular regulation of floral volatile biosynthesis for Lilium cultivars. In an initial pursuit to provide breeders with molecular markers for floral volatile biosynthesis, a total of five commercially available oriental and oriental-trumpet hybrid Lilium cultivars were selected for analytical characterization of floral volatile emission. In total, 66 volatile molecules were qualified and quantitated among all cultivars. Chemical classes of identified volatiles include monoterpene hydrocarbons, monoterpene alcohols and aldehydes, phenylpropanoids, benzenoids, fatty-acid-derived, nitrogen-containing, and amino-acid-derived compounds. In general, the floral volatile profiles of the three oriental-trumpet hybrids were dominated by monoterpene hydrocarbons, monoterpene alcohols and aldehydes, while the two oriental hybrids were dominated by monoterpene alcohols and aldehydes and phenylpropanoids, respectively. Tepal tissues (two petal whirls) emitted the vast majority of total volatile molecules compared to the reproductive organs of the flowers. Tepal volatile profiles were cultivar specific with a high degree of distinction, which indicates the five cultivars chosen will provide an excellent differential genetic environment for gene discovery through comparative transcriptomics in the future. Cloning and assaying transcript accumulation from four floral volatile biosynthetic candidates provided few immediate or obvious trends with floral volatile emission.

*Summary Petunia flower petals emit large amounts of isoeugenol, which has been shown to be synthesized by isoeugenol synthase (PhIGS1) from an ester of coniferyl alcohol, hypothesized to be coniferyl acetate. This paper describes the identification and characterization of a novel petunia gene encoding an enzyme belonging to the BAHD acyltransferase family whose expression correlates with isoeugenol biosynthesis. RNAi suppression of this gene results in inhibition of isoeugenol biosynthesis. Biochemical characterization of the protein encoded by this gene showed that it has acetyltransferase activity and is most efficient with coniferyl alcohol among the alcohol substrates tested. Overall, these data support the conclusion that coniferyl acetate is the substrate of isoeugenol synthase.

In the past decade researchers have reported increasingly comprehensive cDNA libraries of genes associated with floral senescence in floral crops. Analysis of the function of these genes in flower senescence has depended on the slow process of stable transformation and regeneration of the plants. We have adopted an alternative strategy, using virus-induced gene silencing (VIGS) to evaluate the effects on flower senescence of silencing senescence-associated gene. Using purple flowered petunia as our test organism, we infect with a construct where the RNA2 of tobacco rattle virus (TRV) is modified by insertion of chalcone synthase (CHS, a key enzyme in anthocyanin biosynthesis). When virus infection occurs, the plant’s silencing mechanism silences CHS, resulting in white sectors or flowers. Infection with TRV containing a tandem construct of phytoene desaturase (PDS) and CHS resulted in characteristic photobleaching of leaves (due to impaired carotene synthesis), and white patterns on the flowers. Transcripts of CHS and PDS were reduced both in leaves and in flowers confirming simultaneous silencing of both genes by the tandem construct. We tested the effects of infection with TRV containing CHS and a fragment of a petunia gene encoding for 1-aminocyclopropane-1-carboxylate oxidase (ACO). Whether the flowers were treated with ACC or pollinated, the white (silenced) sectors produced less ethylene and senesced later than the purple sectors. These results indicate the value of VIGS with tandem constructs containing CHS as reporter and a target gene as a tool for examining the function of genes isolated from senescing floral tissues.

In many flowering plants, such as petunia (Petunia × hybrida), ethylene produced in floral organs after pollination elicits a series of physiological and biochemical events, ultimately leading to senescence of petals and successful fertilization. Here, we demonstrate, using transgenic ethylene insensitive (44568) and Mitchell Diploid petunias, that multiple components of emission of volatile organic compounds (VOCs) are regulated by ethylene. Expression of benzoic acid/salicylic acid carboxyl methyltransferase (PhBSMT1 and 2) mRNA is temporally and spatially down-regulated in floral organs in a manner consistent with current models for postpollination ethylene synthesis in petunia corollas. Emission of methylbenzoate and other VOCs after pollination and exogenous ethylene treatment parallels a reduction in PhBSMT1 and 2 mRNA levels. Under cyclic light conditions (day/night), PhBSMT mRNA levels are rhythmic and precede emission of methylbenzoate by approximately 6 h. When shifted into constant dark or light conditions, PhBSMT mRNA levels and subsequent methylbenzoate emission correspondingly decrease or increase to minimum or maximum levels observed during normal conditions, thus suggesting that light may be a more critical influence on cyclic emission of methylbenzoate than a circadian clock. Transgenic PhBSMT RNAi flowers with reduced PhBSMT mRNA levels show a 75% to 99% decrease in methylbenzoate emission, with minimal changes in other petunia VOCs. These results implicate PhBSMT1 and 2 as genes responsible for synthesis of methylbenzoate in petunia.

Carotenoids and carotenoid cleavage products play an important and integral role in plant development. The Decreased apical dominance1 (Dad1)/PhCCD8 gene of petunia (Petunia hybrida) encodes a hypothetical carotenoid cleavage dioxygenase (CCD) and ortholog of the MORE AXILLARY GROWTH4 (MAX4)/AtCCD8 gene. The dad1-1 mutant allele was inactivated by insertion of an unusual transposon (Dad-one transposon), and the dad1-3 allele is a revertant allele of dad1-1. Consistent with its role in producing a graft-transmissible compound that can alter branching, the Dad1/PhCCD8 gene is expressed in root and shoot tissue. This expression is upregulated in the stems of the dad1-1, dad2, and dad3 increased branching mutants, indicating feedback regulation of the gene in this tissue. However, this feedback regulation does not affect the root expression of Dad1/PhCCD8. Overexpression of Dad1/PhCCD8 in the dad1-1 mutant complemented the mutant phenotype, and RNA interference in the wild type resulted in an increased branching phenotype. Other differences in phenotype associated with the loss of Dad1/PhCCD8 function included altered timing of axillary meristem development, delayed leaf senescence, smaller flowers, reduced internode length, and reduced root growth. These data indicate that the substrate(s) and/or product(s) of the Dad1/PhCCD8 enzyme are mobile signal molecules with diverse roles in plant development.

Virus-induced gene silencing has been suggested as a powerful and rapid technique for analysis of gene function in plant growth and development (Baulcombe, 1999). Infecting Nicotiana benthamiana with Tobacco Rattle Virus (TRV) containing a fragment of the gene encoding phytoene desaturase, an essential enzyme in carotene synthesis, results in a photo-bleached phenotype in photosynthetic tissues. TRV has several interesting properties that could be utilized in studies of the function of candidate genes in floral senescence; it infects floral tissues, has only mild symptoms, and has a broad host range. We tested the effect of TRV containing PDS fragments on a range of host plants. In most cases, infection resulted in no apparent phenotype or local inoculation effects. In petunia, after a lag of several weeks, the characteristic photo-bleaching of upper portions of the plant indicated movement and silencing of the virus and of PDS. In preliminary tests with a TRV/CHS construct, the typical effects of post-transcriptional gene silencing of CHS in petunia – symmetrical and asymmetrical white patches on the dark purple petals – indicated the ability to use the system in petunia studies.

The plant hormone ethylene regulates many aspects of growth and development. Loss-of-function mutations in ETHYLENE INSENSITIVE2 (EIN2) result in ethylene insensitivity in Arabidopsis, indicating an essential role of EIN2 in ethylene signaling. However, little is known about the role of EIN2 in species other than Arabidopsis. To gain a better understanding of EIN2, a petunia (Petunia × hybrida cv Mitchell Diploid [MD]) homolog of the Arabidopsis EIN2 gene (PhEIN2) was isolated, and the role of PhEIN2 was analyzed in a wide range of plant responses to ethylene, many that do not occur in Arabidopsis. PhEIN2 mRNA was present at varying levels in tissues examined, and the PhEIN2 expression decreased after ethylene treatment in petals. These results indicate that expression of PhEIN2 mRNA is spatially and temporally regulated in petunia during plant development. Transgenic petunia plants with reduced PhEIN2 expression were compared to wild-type MD and ethylene-insensitive petunia plants expressing the Arabidopsis etr1-1 gene for several physiological processes. Both PhEIN2 and etr1-1 transgenic plants exhibited significant delays in flower senescence and fruit ripening, inhibited adventitious root and seedling root hair formation, premature death, and increased hypocotyl length in seedling ethylene response assays compared to MD. Moderate or strong levels of reduction in ethylene sensitivity were achieved with expression of both etr1-1 and PhEIN2 transgenes, as measured by downstream expression of PhEIL1. These results demonstrate that PhEIN2 mediates ethylene signals in a wide range of physiological processes and also indicate the central role of EIN2 in ethylene signal transduction.

Pollen viability, in-vivo pollen tube growth, fruit ripening, seed germination, seed weight, whole plant vigor, and natural fl ower senescence were investigated in homozygous and heterozygous transgenic ethylene-insensitive CaMV35S::etr1-1 petunias (Petunia ×hybrida 'Mitchell Diploidʼ). Homozygous or heterozygous plants were used to determine any maternal and/or paternal effects of the CaMV35S::etr1-1 transgene. All experiments except for those used to determine natural fl ower senescence characteristics were conducted in both high and low temperature greenhouses to determine the effect of temperature stress on transgenic plants when compared to wild-type. Results indicated that ethylene-insensitive plants had a decrease in pollen viability, root dry mass, seed weight, and seed germination. Fruit ripening, seed germination, and seed weight were maternally regulated. In contrast, the CaMV35S::etr1-1 transgene is completely dominant in its effect on natural fl ower senescence. The plant hormone ethylene is involved in many physiologi- cal processes in plants including fruit ripening, petal senescence, abscission, and seed germination (Abeles et al., 1992). The senes- cence of many fl owers is accompanied by a dramatic increase in ethylene production resulting in petal wilting, abscission or color fading. Treatment of ethylene-sensitive fl owers with chemical inhibitors of ethylene biosynthesis or perception reduces this burst of ethylene production and delays visual symptoms of corolla senescence (Jones and Woodson, 1997; Serek et al., 1995; White- head et al., 1984). Genetic modifi cation of ethylene biosynthetic and signal transduction pathways has provided further evidence of ethyleneʼs role in regulating fl ower senescence (Bovy et al., 1999; Savin et al., 1995; Wilkinson et al., 1997). Since many economically important fl oral crops are grown for their display, the control of ethylene perception is essential for increasing vase life and enhancing quality. Since one of the major concerns in the fl oriculture industry is fl ower longevity, research has been conducted to use genetic engineering on fl oriculture crops that are insensitive to ethylene. Wilkinson et al. (1997) transformed petunia with a dominant mu- tant Arabidopsis ethylene receptor, etr1-1, under the control of a constitutive Caulifl ower Mosaic Virus 35S (CaMV35S) promoter to produce ethylene-insensitivity throughout the whole plant. These petunias (44568) had an increase in natural and pollina- tion-induced fl ower longevity compared to wild type 'Mitchell Diploidʼ (MD) plants, but had physiological side effects that limit their commercial use (Gubrium et al., 2000; Wilkinson et al., 1997). Ethylene-insensitive petunias and tomatoes both showed a signifi cant reduction in adventitious root formation, and even exogenous treatments with auxin did not increase adventitious root formation to the level of untreated wild-type controls (Clark

Cytokinins have been shown to delay the onset of leaf senescence. The focus of this project was to produce transgenic petunia (Petunia ×hybrida) plants that over-produced endogenous cytokinins in a senescence specifi c manner. This was achieved by transforming plants with the IPT (isopentenyl transferase) gene driven by the senescence-associated transcriptional promoter, PSAG12. Two independent transgenic events produced T1 and T2 generation seedling lines that demonstrated the desired nonsenescent phenotype in progeny trials. These lines were used to evaluate the horticultural performance of PSAG12-IPT petunia plants in terms of delayed senescence, rooting of vegetative cuttings, lateral branch growth, fl ower number, fl oral timing, and fruit set. Although both lines displayed a delayed senescence phenotype the two PSAG12-IPT transgenic lines differed from each other in regard to other horticultural traits. In addition to delayed leaf senescence, line I-1-7 also demonstrated a decrease in adventitious rooting and an increased number of branches during plant production. Line I-3-18 also demonstrated a delayed leaf senescence phenotype; however, plants of this line were not greatly altered in any other horticultural performance traits in comparison to wild-type 'V26ʼ. IPT transcript was detected in young fully expanded leaves of both lines, although mRNA levels were higher in I-1-7 plants. A greater than 50-fold increase in IPT transcript abundance was detected in leaves of transgenic plants following drought stress. These results demonstrate that it is possible to use PSAG12-IPT to produce transgenic plants with delayed leaf senescence but differences in plant morphology between transgenic lines exist, which may alter horticultural performance characteristics. Senescence is the fi nal developmental process in the lifecycle of a leaf. It is a process through which the nutrients contained within the macromolecules (e.g., chlorophyll, proteins, nucleic acids) of leaf cells are metabolized to basic components and trans- ported to the growing shoot and reproductive organs of the plant. This strategy imparts a survival advantage for plants that grow under adverse conditions where nutrients are limiting (Nooden, 1988). Natural leaf senescence in many plants is characterized by yellowing or chlorosis of the lower leaves as nutrients and other components of the cells are degraded (esp. chlorophyll). In horticultural terms, chlorosis can decrease the aesthetic appear- ance and thereby decrease the salability of that plant. With so much at risk, it is no wonder that growers and researchers alike have tried to understand the components of leaf senescence in order to prevent it. One way to prevent leaf senescence is through the manipulation of cytokinins. Cytokinins are an important class of plant hormones that infl uence numerous aspects of plant growth and development. They have been shown to delay and, in some cases, reverse the leaf senescence process (Gan and Amasino, 1996). There is an inverse correlation between the concentration of cytokinins in leaves and the magnitude of leaf senescence. This can be dramati- cally seen as the formation of green islands on leaves infected by cytokinin-producing pathogens, such as Helminthosporium teres (Angra and Mandahar, 1991). The use of applied cytokinins to prevent senescence is limited because it is diffi cult to supply them

The molecular mechanisms responsible for postpollination changes in floral scent emission were investigated in snapdragon cv Maryland True Pink and petunia cv Mitchell flowers using a volatile ester, methylbenzoate, one of the major scent compounds emitted by these flowers, as an example. In both species, a 70 to 75% pollination-induced decrease in methylbenzoate emission begins only after pollen tubes reach the ovary, a process that takes between 35 and 40 h in snapdragon and approximately 32 h in petunia. This postpollination decrease in emission is not triggered by pollen deposition on the stigma. Petunia and snapdragon both synthesize methylbenzoate from benzoic acid and S-adenosyl-l-methionine (SAM); however, they use different mechanisms to downregulate its production after pollination. In petunia, expression of the gene responsible for methylbenzoate synthesis is suppressed by ethylene. In snapdragon, the decrease in methylbenzoate emission is the result of a decrease in both S-adenosyl-l-methionine:benzoic acid carboxyl methyltransferase (BAMT) activity and the ratio of SAM to S-adenosyl-l-homocysteine ("methylation index") after pollination, although the BAMT gene also is sensitive to ethylene.

The delayed leaf senescence process has been engineered in transgenic Petunia x hybrida plants containing the homeobox knotted-1 gene (kn1), a mutant maize gene that is thought to be involved in cytokinin metabolism. Petunia plants were transformed with the kn1 gene fused to a senescence-associated promoter (SAG 12 ) using Agrobacterium. Putative P SAG12 -kn1 lines were subjected to drought stress treatments, and leaf senescence was evaluated in the To and T 1 generations. A wide range of delayed senescence phenotypes was observed, indicating that the promoter SAG 12 did not always present complete regulation. Although expression of the kn1 transgene delayed leaf senescence, it also caused some morphological and physiological alterations, which ranged from mild to severe in their intensity. Most of the plants with a non-senescent phenotype showed only mild phenotypic alterations, which were not considered detrimental to plant performance. Along with the trait of delayed leaf senescence, plants with mild phenotypes had a more compact architecture, whereas plants with a severe phenotype were stunted in growth. Our data support the concept that the P SAG12 -kn1 genetic construct can be used to delay leaf senescence, but the mechanism(s) by which kn1 gene affects the senescence process remains to be elucidated.

Uniconazole was applied as a spray to the surface of container media prior to planting bedding plant plugs. This medium spray was compared to a standard whole-plant spray applied 2 weeks after planting. For petunia (Petunia ×hybrida Vilm.) and coleus (Solenostemon scutellarioides L.) the efficacy of the medium spray was similar to the whole-plant spray. However, for impatiens (Impatiens wallerana Hook. f.) and vinca [Catharanthus roseus (L.) G. Don.] the medium spray had greater efficacy than the whole-plant spray. Increased concentrations of uniconazole in the medium spray decreased plant height; however, the effect of higher concentrations was greater in a medium with out pine bark compared to a medium with pine bark as a component. In the above experiments, uniconazole was applied in a volume of 200 mL·m-2. In a test where spray volume varied, there was a negative linear relationship between plant height and spray volume. Chemical name used: (E)-(+)-(S)-1-(4-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)-pent-1-ane-3-ol (uniconazole).

Breeding for a subjective goal such as flavor is challenging, as many blueberry cultivars are grown worldwide, and identifying breeding targets relating to blueberry flavor biochemistry that have a high degree of genetic control and low environmental variability are priorities. A variety of biochemical compounds and physical characters induce the sensory responses of taste, olfaction, and somatosensation, all of which interact to create what is perceived flavor. The goal of this study was to identify the flavor compounds with a larger genetic versus environmental component regulating their expression over an array of cultivars, locations, and years. Over the course of three years, consumer panelists rated overall liking, texture, sweetness, sourness, and flavor intensity of 19 southern highbush blueberry (Vaccinium corymbosum hybrids) genotypes in 30 sensory panels. Significant positive correlations to overall liking of blueberry fruit (P

Chlorophyll (Chl) and total soluble protein decreased and proteolytic activity increased over a 12-day period during dark-induced senescence in detached leaves of Tara, a yellowing cultivar (Y) of Dendranthema grandiflora. In Boaldi, a non-yellowing cultivar (NY), Chl and soluble protein remained near initial levels and little change in proteolytic activity was observed. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of soluble proteins showed no major differences in banding patterns between the two cultivars at day 0; however, all of the resolved proteins were diminished in Tara by day 12. On the other hand, in NY Boaldi, the intensity of the protein bands did not change over the 12-day period. Attached and detached leaves exhibited similar senescence patterns for each cultivar. Ethylene (100 μl l -1 ) accelerated the rate of Chl loss in detached leaves of Tara, but had no effect on Boaldi. These observations suggest that Boaldi is a stay-green genotype, possibly a functional type. The results are discussed in relation to the role of ethylene in chrysanthemum leaf senescence.

ADDITIONAL INDEX WORDS. ACC (1aminocyclopropane-1-carboxylic acid), ethylene biosynthesis, flower petal abscission, pollination, seedling triple response SUMMARY. Experiments were conducted to determine if the seedling hypocotyl elongation and petal abscission assays could be used to identify differences in ethylene sensitivity among seedling geranium (Pelargonium ×hortorum) cultivars. When seedlings of six geranium cultivars were germinated and grown in the dark in the presence of the ethylene biosynthetic precursor 1-aminocyclopropane1-carboxylic acid (ACC) at various concentrations, they exhibited the triple response (measured as reduced hypocotyl length). While seedlings from all six cultivars were sensitive to ACC, ‘Scarlet Elite’ seedlings were most sensitive, and ‘Multibloom Lavender’, ‘Elite White’ and ‘Ringo 2000 Salmon’ seedlings were the least sensitive when germinated and grown on 20 mM [2022 mg·L–1 (ppm)] ACC. Florets representing three developmental stages of each of the six cultivars were exposed to 1 μL·L–1 of exogenous ethylene for 0, 30, or 60 min to determine if differences in cultivar sensitivity could be determined for petal abscission. Of the six cultivars tested, ‘Ringo 2000 Salmon’, ‘Multibloom Lavender’ and ‘Elite White’ were the least ethylene sensitive. Florets were also self-pollinated to test for cultivar differences in ethylene synthesis and subsequent petal abscission. Ethylene production and petal abscission were both promoted in selfpollinated florets compared to nonpollinated florets. ‘Ringo 2000 Salmon’, ‘Multibloom Lavender’ and ‘Elite White’ florets produced similar amounts of ethylene as all other cultivars, but abscised fewer petals after pollination. Our results indicate that the seedling hypocotyls elongation assay may be used to identify geranium cultivars with reduced sensitivity to ethylene. The data also suggest that genetic variability exists among geraniums for both ethylene sensitivity and biosynthesis. Ethylene induces premature floral abscission in numerous plant species (Abeles et al., 1992). Diploid zonal geraniums are among the most sensitive species with regard to ethylene-induced petal abscission; exposure of florets to 1 μL·L–1 ethylene can result in complete abscission in as little as 1 h (Evensen et al., 1993; Clark et al., 1997). Exposure to trace amounts of ethylene during postharvest transit and marketing can cause petal drop and significantly reduce the visual quality of geraniums (Armitage et al., 1980) and other important floriculture crops. Additionally, ethylene plays a role in the rapid abscission of the corolla following pollination (Abeles et al., 1992). A rapid increase in ethylene production by floral tissues after pollination, followed by accelerated corolla senescence and abscission, was reported for carnation (Dianthus caryophyllus) (Nichols, 1977), digitalis (Digitalis purpurea) (Stead and Moore, 1979, 1983), petunia (Petunia hybrida) (Pech et al., 1987; Whitehead et al., 1984), and geranium (Clark et al., 1997). Petal abscission by pollinated geranium florets is completely but reversibly blocked by 2,5 norbornadiene, an ethylene action inhibitor, indicating that endogenous ethylene perception is essential for the abscission response (Clark et al., 1997). Ethylene also has a regulatory effect on the growth of seedlings following germination. Dark-grown etiolated pea (Pisum sativum) seedlings exposed to exogenous ethylene exhibited: 1) decreased elongation of the hypocotyl (Arshad and Frankenburger, 1988; Koch and Moore, 1990); 2) increased hypocotyl diameter; and 3) exaggerated plumular hooks (Goeschl et al., 1966). This triple response of seedlings was used to identify ethyleneinsensitive mutants as a means to elucidate the mechanism of ethylene perception and action at the molecular level in arabidopsis (Arabidopsis thaliana) (Guzman and Ecker, 1990; Harpam et al., 1991) and tomato (Lycopersicon esculentum) (Lanahan et al., 1994). The effect of ethylene on hypocotyl elongation appears to be concentration-dependent; as exogenous ethylene concentration is increased, hypocotyl length is decreased up to a point of saturation (Goeschl and Kays, 1975; Harpham et al., 1991). Zonal geranium cultivars differ Environmental Horticulture Dept., University of Florida, Gainesville, FL 32611-0670.

A series of experiments on ethylene-insensitive (EI) petunia plants (Petunia ×hybrida Hort. Vilm.-Andr.) generated in two genetic backgrounds were conducted to determine the involvement of ethylene in horticultural performance. Experiments examined various aspects of horticultural performance: days to flower, flower senescence after pollination and without pollination, fruit set and ripening, and adventitious root formation on vegetative stem cuttings. The development of EI plants was altered in several ways. Time from seed sowing to first flower anthesis was decreased by a week for EI plants grown at 26/21 °C. Flower senescence in nonpollinated and self-pollinated flowers was delayed in all EI plants compared to wild-type plants. Fruit set percentage on EI plants was slightly lower than on wild-type plants and fruit ripening on EI plants was delayed by up to 7 days. EI plants produced fewer commercially acceptable rooted cuttings than wild-type plants. There was a basic difference in the horticultural performance of the two EI lines examined due to a difference in the genetic backgrounds used to generate the lines. EI plants displayed better horticultural performance when grown with day/night temperatures of 26/21 °C than 30/24 °C. These results suggest that tissue-specific ethylene insensitivity as well as careful consideration of the genetic background used in transformation procedures and growth conditions of etr1-1 plants will be required to produce commercially viable transgenic floriculture crops. EI petunias provide an ideal model system for studying the role of ethylene in regulating various aspects of plant reproduction.

We have isolated two cDNAs from geranium, PhETR1 and PhETR2. The deduced amino acid sequences of PhETR1 and PhETR2 share 78% and 79% identity with ETR1 from Arabidopsis thaliana respectively. These genes are members of a multigene family and are expressed at moderate levels in leaves, pedicels, sepals, pistils and petals, and at very low levels in roots. PhETR1 and PhETR2 mRNAs are expressed in geranium florets long before they are receptive to pollination and transcript levels remain constant throughout floral development. Message levels of PhETR1 and PhETR2 in pistils and receptacles are unaffected by self-pollination or treatment with 1 μ/l ethylene that induces petal abscission. Our results indicate that the amount of PhETR1 and PHETR2 mRNA is not indicative of the level of sensitivity of geranium florets to ethylene.