Mahmoud Wahb-Allah

Development of a Cucumis sativus TILLinG Platform for Forward and Reverse Genetics

Background Cucumber (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 Findings A 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. Conclusions We 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.

Conocarpus Biochar Induces Changes in Soil Nutrient Availability and Tomato Growth Under Saline Irrigation

Thermally modified organic materials commonly known as biochar have gained popularity of being used as a soil amendment. Little information, however, is available on the role of biochar in alleviating the negative impacts of saline water on soil productivity and plant growth. This study, therefore, was conducted to investigate the effects of Conocarpus biochar (BC) and organic farm residues (FR) at different application rates of 0.0% (control), 4.0% and 8.0% (weight/weight) on yield and quality of tomatoes grown on a sandy soil under drip irrigation with saline or non-saline water. The availability of P, K, Fe, Mn, Zn and Cu to plants was also investigated. The results demonstrated clearly that addition of BC or FR increased the vegetative growth, yield and quality parameters in all irrigation treatments. It was found that salt stress adversely affected soil productivity, as indicated by the lower vegetative growth and yield components of tomato plants. However, this suppressing effect on the vegetative growth and yield tended to decline with application of FR or BC, especially at the high application rate and in the presence of biochar. Under saline irrigation system, for instance, the total tomato yield increased over the control by 14.0%–43.3% with BC and by 3.9%–35.6% with FR. These could be attributed to enhancement effects of FR or BC on soil properties, as indicated by increases in soil organic matter content and nutrient availability. Therefore, biochar may be effectively used as a soil amendment for enhancing the productivity of salt-affected sandy soils under arid conditions.

Characterisation of alleles of tomato light signalling genes generated by TILLING

Targeting Induced Local Lesions IN Genomes (TILLING) combines chemical mutagenesis with high throughput screening to allow the generation of alleles of selected genes. In this study, TILLING has been applied to produce a series of mutations in genes encoding essential components of the tomato light signal transduction pathway in an attempt to enhance fruit nutritional quality. Point mutations to DEETIOLATED1 (DET1), which is responsible for the high pigment2 (hp2) tomato mutant, resulted in elevated levels of both carotenoid and phenylpropanoid phytonutrients in ripe fruit, whilst immature fruit showed increased chlorophyll content, photosynthetic capacity and altered fruit morphology. Furthermore, genotypes with mutations to the UV-DAMAGED DNA BINDING PROTEIN 1 (DDB1), COP1 and COP1like were also characterised. These genotypes largely did not display phenotypes characteristic of mutation to light signalling components but their characterisation has enabled interrogation of structure function relationships of the mutated genes.