BackgroundFruit ripening and softening are key traits that have an effect on food supply, fruit nutritional value and consequently, human health. Since ethylene induces ripening of climacteric fruit, it is one of the main targets to control fruit over ripening that leads to fruit softening and deterioration. The characterization of the ethylene pathway in Arabidopsis and tomato identified key genes that control fruit ripening.Methodology/Principal FindingsTo engineer melon fruit with improved shelf-life, we conducted a translational research experiment. We set up a TILLING platform in a monoecious and climacteric melon line, cloned genes that control ethylene production and screened for induced mutations that lead to fruits with enhanced shelf life. Two missense mutations, L124F and G194D, of the ethylene biosynthetic enzyme, ACC oxidase 1, were identified and the mutant plants were characterized with respect to fruit maturation. The L124F mutation is a conservative mutation occurring away from the enzyme active site and thus was predicted to not affect ethylene production and thus fruit ripening. In contrast, G194D modification occurs in a highly conserved amino acid position predicted, by crystallographic analysis, to affect the enzymatic activity. Phenotypic analysis of the G194D mutant fruit showed complete delayed ripening and yellowing with improved shelf life and, as predicted, the L124F mutation did not have an effect.Conclusions/SignificanceWe constructed a mutant collection of 4023 melon M2 families. Based on the TILLING of 11 genes, we calculated the overall mutation rate of one mutation every 573 kb and identified 8 alleles per tilled kilobase. We also identified a TILLING mutant with enhanced fruit shelf life. This work demonstrates the effectiveness of TILLING as a reverse genetics tool to improve crop species. As cucurbits are model species in different areas of plant biology, we anticipate that the developed tool will be widely exploited by the scientific community.

Cooling greenhouses is essential to provide a suitable environment for plant growth in arid regions characterized by brackish water resources. However, using conventional cooling methods are facing many challenges. Filtering out near infra-red radiation (NIR) at the greenhouse cover can significantly reduce the heating load and can solve the overheating problem of the greenhouse air. This paper is to review (i) the problems of using conventional cooling methods and (ii) the advantages of greenhouse covers that incorporate NIR reflectors. This survey focuses on how the cover type affects the transmittance of photosynthetically active radiation (PAR), the reflectance or absorptance of NIR and the greenhouse air temperature. NIR-reflecting plastic films seem to be the most suitable, low cost and simple cover for greenhouses under arid conditions. Therefore, this review discusses how various additives should be incorporated in plastic film to increase its mechanical properties, durability and ability to stand up to extremely harsh weather. Presently, NIR-reflecting covers are able to reduce greenhouse air temperature by no more than 5°C. This reduction is not enough in regions where the ambient temperature may exceed 45°C in summer. There is a need to develop improved NIR-reflecting plastic film covers.

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Soil microorganisms might be assessed for their capabilities of plant growth promotion in order to identify heat tolerant strategies for crop production. The planned study was conducted to determine the potential of heat tolerant plant growth promoting rhizobacteria (PGPR) in mitigating heat stress effects in tomato. Bacillus cereus was evaluated for plant growth promoting activities and assessed for 1-aminocyclopropane-1-carboxylate (ACC-deaminase) (0.76–C0.9 μM/mg protein/h), and exopolysaccharide (0.66–C0.91 mg/mL) under normal and heat stressed conditions. Plant growth regulators were evaluated through High Performance Liquid Chromatography. Bacterial inoculation effects on important physiological and biochemical parameters were evaluated under normal and heat stressed conditions in growth chamber. The morphological-physiological traits significantly revealed drastic effects on both of un-inoculated tomato varieties under heat stress conditions. Bacterial augmentation significantly promoted shoot, root length, leaf surface area, fresh and dry weight. Heat stress enhanced extracellular polymeric substances (EPS) production and cleavage of ACC into a-ketobutyrate and ammonia due to ACC-deaminase producing bacteria that significantly reduced the adverse effects of heat on tomato growth. In conclusion, the applied plant growth promoting rhizobacteria (PGPR) bacterial strain proved as potential candidate for improving tomato crop growing under heat stressed conditions. However, it is highly suggested to validate the current results by conducting field trials.

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BackgroundCucumber (Cucumis sativus) belongs to the Cucurbitaceae family that includes more than 800 species. The cucumber genome has been recently sequenced and annotated. Transcriptomics and genome sequencing of many plant genomes are providing information on candidate genes potentially related to agronomically important traits. To accelerate functional characterization of these genes in cucumber we have generated an EMS mutant population that can be used as a TILLinG platform for reverse genetics.Principal FindingsA population of 3,331 M2 mutant seed families was generated using two EMS concentrations (0.5% and 0.75%). Genomic DNA was extracted from M2 families and eight-fold pooled for mutation detection by ENDO1 nuclease. To assess the quality of the mutant collection, we screened for induced mutations in five genes and identified 26 mutations. The average mutation rate was calculated as 1/1147 Kb giving rise to approximately 320 mutations per genome. We focused our characterization on three missense mutations, G33C, S238F and S249F identified in the CsACS2 sex determination gene. Protein modeling and crystallography studies predicted that mutation at G33 may affect the protein function, whereas mutations at S238 and S249 may not impair the protein function. As predicted, detailed phenotypic evaluation showed that the S238F and the S249F mutant lines had no sexual phenotype. In contrast, plants homozygous for the G33C mutation showed a complete sexual transition from monoecy to andromonoecy. This result demonstrates that TILLinG is a valuable tool for functional validation of gene function in crops recalcitrant to transgenic transformation.ConclusionsWe have developed a cucumber mutant population that can be used as an efficient reverse genetics tool. The cucumber TILLinG collection as well as the previously described melon TILLinG collection will prove to be a valuable resource for both fundamental research and the identification of agronomically-important genes for crop improvement in cucurbits in general.

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