Diversity arrays technology (DArT) markers in apple for genetic linkage maps

Diversity Arrays Technology (DArT) provides a high-throughput whole-genome genotyping platform for the detection and scoring of hundreds of polymorphic loci without any need for prior sequence information. The work presented here details the development and performance of a DArT genotyping array for apple. This is the first paper on DArT in horticultural trees. Genetic mapping of DArT markers in two mapping populations and their integration with other marker types showed that DArT is a powerful high-throughput method for obtaining accurate and reproducible marker data, despite the low cost per data point. This method appears to be suitable for aligning the genetic maps of different segregating populations. The standard complexity reduction method, based on the methylation-sensitive PstI restriction enzyme, resulted in a high frequency of markers, although there was 52–54% redundancy due to the repeated sampling of highly similar sequences. Sequencing of the marker clones showed that they are significantly enriched for low-copy, genic regions. The genome coverage using the standard method was 55–76%. For improved genome coverage, an alternative complexity reduction method was examined, which resulted in less redundancy and additional segregating markers. The DArT markers proved to be of high quality and were very suitable for genetic mapping at low cost for the apple, providing moderate genome coverage

Identification and Functional Characterization of the Lactococcus lactis rfb Operon, Required for dTDP-Rhamnose Biosynthesis

dTDP-rhamnose is an important precursor of cell wall polysaccharides and rhamnose-containing exopolysaccharides (EPS) in Lactococcus lactis. We cloned the rfbACBD operon from L. lactis MG1363, which comprises four genes involved in dTDP-rhamnose biosynthesis. When expressed in Escherichia coli, the lactococcal rfbACBD genes could sustain heterologous production of the Shigella flexneri O antigen, providing evidence of their functionality. Overproduction of the RfbAC proteins in L. lactis resulted in doubled dTDP-rhamnose levels, indicating that the endogenous RfbAC activities control the intracellular dTDP-rhamnose biosynthesis rate. However, RfbAC overproduction did not affect rhamnose-containing B40-EPS production levels. A nisin-controlled conditional RfbBD mutant was unable to grow in media lacking the inducer nisin, indicating that the rfb genes have an essential role in L. lactis. Limitation of RfbBD activities resulted in the production of altered EPS. The monomeric sugar of the altered EPS consisted of glucose, galactose, and rhamnose at a molar ratio of 1:0.3:0.2, which is clearly different from the ratio in the native sugar. Biophysical analysis revealed a fourfold-greater molecular mass and a twofold-smaller radius of gyration for the altered EPS, indicating that these EPS are more flexible polymers with changed viscosifying properties. This is the first indication that enzyme activity at the level of central carbohydrate metabolism affects EPS composition

Fine mapping of the gene Rvi18 (V25) for broad-spectrum resistance to apple scab, and development of a linked SSR marker suitable for marker-assisted breeding

Apple scab, caused by the fungal pathogen Venturia inaequalis, is one of the most devastating diseases for the apple growing industry in temperate zones with humid springs and summers. Breeding programs around the world have identified several sources of resistance, of which the Rvi6 (Vf) gene from Malus floribunda 821 has been the most widely used. The appearance of Rvi6-virulent strains of V. inaequalis in several European countries have underlined the necessity of pyramiding different effective resistance genes for durably resistant cultivars. Here we report the mapping of the new apple scab resistance gene Rvi18 (V25) from the selection 1980-015-025 of the apple breeding program at Wageningen University and Research Centre, The Netherlands. This gene was fine mapped on the proximal part of LG11 to a region of 34 Kb in the apple genome sequence of ‘Golden Delicious’, using 894 progeny plants, and SSR, DArT, AFLP, and SNP markers. One gene on the ‘Golden Delicious’ reference genome was identified as the potential susceptibility allele of the resistance gene. Moreover, an SSR marker has been developed of which one of its amplicons sizes is highly specific for Rvi18, thus facilitating the directed pyramiding of resistance genes through marker assisted breeding

Solyntus, the New Highly Contiguous Reference Genome for Potato (Solanum tuberosum)

With the rapid expansion of the application of genomics and sequencing in plant breeding, there is a constant drive for better reference genomes. In potato (Solanum tuberosum), the third largest food crop in the world, the related species S. phureja, designated “DM”, has been used as the most popular reference genome for the last 10 years. Here, we introduce the de novo sequenced genome of Solyntus as the next standard reference in potato genome studies. A true Solanum tuberosum made up of 116 contigs that is also highly homozygous, diploid, vigorous and self-compatible, Solyntus provides a more direct and contiguous reference then ever before available. It was constructed by sequencing with state-of-the-art long and short read technology and assembled with Canu. The 116 contigs were assembled into scaffolds to form each pseudochromosome, with three contigs to 17 contigs per chromosome. This assembly contains 93.7% of the single-copy gene orthologs from the Solanaceae set and has an N50 of 63.7 Mbp. The genome and related files can be found at https://www.plantbreeding.wur.nl/Solyntus/. With the release of this research line and its draft genome we anticipate many exciting developments in (diploid) potato research

Exploration of a Resequenced Tomato Core Collection for Phenotypic and Genotypic Variation in Plant Growth and Fruit Quality Traits

A tomato core collection consisting of 122 gene bank accessions, including landraces, old cultivars, and wild relatives, was explored for variation in several plant growth, yield and fruit quality traits. The resequenced accessions were also genotyped with respect to a number of mutations or variations in key genes known to underlie these traits. The yield-related traits fruit number and fruit weight were much higher in cultivated varieties when compared to wild accessions, while, in wild tomato accessions, Brix was higher than in cultivated varieties. Known mutations in fruit size and shape genes could well explain the fruit size variation, and fruit colour variation could be well explained by known mutations in key genes of the carotenoid and flavonoid pathway. The presence and phenotype of several plant architecture affecting mutations, such as self-pruning (sp), compound inflorescence (s), jointless-2 (j-2), and potato leaf (c) were also confirmed. This study provides valuable phenotypic information on important plant growth- and quality-related traits in this collection. The allelic distribution of known genes that underlie these traits provides insight into the role and importance of these genes in tomato domestication and breeding. This resource can be used to support (precision) breeding strategies for tomato crop improvement