SNP GENOTYPING IN TETRAPLOID CUT ROSES

Abstract

Genetic analysis in tetraploid rose is complex: inheritance of genes could be disomic, keeping the two original diploid genomes separate but it could also be tetrasomic, permitting the four homologous chromosomes to pair and recombine during meiosis, and including the possibility of double reduction. Marker-assisted breeding (MAB) is particularly suited for traits such as disease resistance since disease resistance assays are laborious, time and money consuming, but we need to extend the number of markers for better coverage of the genome before being able to find associations and apply MAB. Our goal was to produce hundreds of single nucleotide polymorphism (SNP) markers to provide the coverage needed. Disease resistance, more specifically powdery mildew resistance is our primary target because chemical pest control will be increasingly limited. Molecular markers could provide a good tool to facilitate selection in an earlier stage of a breeding program. We investigated the inheritance of powdery mildew resistance in an F1 population originating from a cross between two tetraploid genotypes, P540 and P867, differing in powdery mildew resistance. SNP marker development started with Next Generation Sequencing (Illumina GAII) of cDNA obtained from RNA isolated from flowers at three developmental stages for each parent. We identified a large number of putative single nucleotide polymorphisms (SNPs) between and within both parents. SNPs that were expected to segregate as simplex × nulliplex and duplex × nulliplex markers were selected for genotyping the entire F1 population using the KASP platform (KBioscience). For a pilot study on a limited number of SNPs we defined specific selection criteria resulting in a set of 20 SNPs for testing on the population. It was found that selecting the SNPs according to the ratio of the number of reads from the Illumina sequencing data did not guarantee the appropriate segregation ratios in the progeny. Continued research will focus on identifying haplotypes and on QTL mapping of powdery mildew resistance. We will present results on the criteria used to select SNP markers, success rate, the realised segregation ratios in the offspring, the reconstruction of the homologous chromosomes (haplotypes) in the cross, and QTL mapping of powdery mildew resistance.