Identification and validation of SNP markers linked to seed toxicity in Jatropha curcas L.

Edible/non-toxic varieties of Jatropha curcas L. are gaining increasing attention, providing both oil as biofuel feedstock or even as edible oil and the seed kernel meal as animal feed ingredient. They are a viable alternative to the limitation posed by the presence of phorbol esters in toxic varieties. Accurate genotyping of toxic/non-toxic accessions is critical to breeding management. The aim of this study was to identify SNP markers linked to seed toxicity in J. curcas. For SNP discovery, NGS technology was used to sequence the whole genomes of a toxic and non-toxic parent along with a bulk of 51 toxic and 30 non-toxic F2 plants. To ascertain the association between SNP markers and seed toxicity trait, candidate SNPs were genotyped on 672 individuals segregating for seed toxicity and two collections of J. curcas composed of 96 individuals each. In silico SNP discovery approaches led to the identification of 64 candidate SNPs discriminating non-toxic and toxic samples. These SNPs were mapped on Chromosome 8 within the Linkage Group 8 previously identified as a genomic region important for phorbol ester biosynthesis. The association study identified two new SNPs, SNP_J22 and SNP_J24 significantly linked to low toxicity with R2 values of 0.75 and 0.54, respectively. Our study released two valuable SNP markers for high-throughput, marker-assisted breeding of seed toxicity in J. curcas.

High-throughput SNP discovery and genotyping in durum wheat (Triticum durum Desf.).

We describe the application of complexity reduction of polymorphic sequences (CRoPS(®)) technology for the discovery of SNP markers in tetraploid durum wheat (Triticum durum Desf.). A next-generation sequencing experiment was carried out on reduced representation libraries obtained from four durum cultivars. SNP validation and minor allele frequency (MAF) estimate were carried out on a panel of 12 cultivars, and the feasibility of genotyping these SNPs in segregating populations was tested using the Illumina Golden Gate (GG) technology. A total of 2,659 SNPs were identified on 1,206 consensus sequences. Among the 768 SNPs that were chosen irrespective of their genomic repetitiveness level and assayed on the Illumina BeadExpress genotyping system, 275 (35.8%) SNPs matched the expected genotypes observed in the SNP discovery phase. MAF data indicated that the overall SNP informativeness was high: a total of 196 (71.3%) SNPs had MAF >0.2, of which 76 (27.6%) showed MAF >0.4. Of these SNPs, 157 were mapped in one of two mapping populations (Meridiano × Claudio and Colosseo × Lloyd) and integrated into a common genetic map. Despite the relatively low genotyping efficiency of the GG assay, the validated CRoPS-derived SNPs showed valuable features for genomics and breeding applications such as a uniform distribution across the wheat genome, a prevailing single-locus codominant nature and a high polymorphism. Here, we report a new set of 275 highly robust genome-wide Triticum SNPs that are readily available for breeding purposes.

Functional differentiation of the sugar beet root system as indicator of developmental phase change.

Developmental phase transitions in the plant root system have not been well characterized. In this study we compared the dynamics of sucrose accumulation with changes in gene expression analyzed with cDNA-amplified fragment length polymorphism (AFLP) in the developing tap root of sugar beet (Beta vulgaris, L.) during the first 9 weeks after emergence (WAE). Although differences between lines were evident as soon as 9 WAE, sucrose showed a marked increase in the rate of accumulation between 4 and 6 WAE and a remarkable shift in gene expression was observed between 5 and 6 WAE. These changes were evident in two unrelated genetic backgrounds and suggest that physiological and gene expression changes represent a functional differentiation of the tap root. These changes were considered as indicators of a developmental change in the sugar beet root system. To identify genes and metabolic pathways involved in this developmental shift, a root cDNA library was hybridized with probes enriched for 3- and 7-WAE transcripts and differentially expressed transcripts were analyzed by cDNA microarray. Several genes involved in the regulation of tissue development were found to be differentially regulated. Genes involved in protein metabolism, disease-related and secretory system were upregulated before the functional differentiation transition, while genes under hormonal control were upregulated after the functional differentiation transition. This developmental phase change of the root system is important to understand plant developmental regulation at the whole-plant level and will likely be useful as early selection parameter in breeding programs.

Fluorometric sucrose evaluation for sugar beet.

Sucrose is the economic product from sugar beet. Disease resistance is often available in low-sucrose genotypes and, prior to the deployment of such novel genes as available into the cultivated spectrum, selection for increased sucrose content is required during introgression. The objective of this work was to evaluate a relatively rapid and inexpensive enzymatic-fluorometric microtiter plate assay for sucrose quantification in sugar beet root dry matter, both for progeny testing in the greenhouse and for evaluation of field-grown mother roots. As determined using HPLC, sucrose content in diverse populations of sugar and table beet assayed over various developmental stages ranged from 0.213 to 2.416 mmol g(-1) of dry matter, and these values were used as references for both refractometry and enzymatic-fluorometric assay. As expected, refractometric analysis generally overestimated sucrose content. Enzymatic-fluorometric analyses were reasonably well correlated with HPLC results for young greenhouse-grown root tissues (R2 = 0.976), and less so with older field-grown roots (R2 = 0.605), for unknown reasons. Enzymatic-fluorometric assays may be best deployed for progeny testing of young seedlings.