Genotyping by Sequencing for SNP-Based Linkage Analysis and the Development of KASPar Markers for Male Sterility and Polyembryony in Citrus

Polyembryony and male sterility (MS) are essential characters for citrus breeding. MS, coupled with parthenocarpy, allows for addressing the diversification of diploid seedless mandarin varieties, and nucleocytoplasmic MS is the most prevalent system. Polyembryony limits the use of seed parents in scion breeding programs, and the recovery of monoembryonic hybrids to be used as female parents is a crucial pre-breeding component. The objectives of this work were the identification of SNPs closely linked with the genes implied in these traits for marker-assisted selection. Genotyping by sequencing was used to genotype 61 diploid hybrids from an F1 progeny recovered from crossing 'Kiyomi' and 'Murcott' tangors. A total of 6444 segregating markers were identified and used to establish the two parental genetic maps. They consisted of 1374 and 697 markers encompassing 1416.287 and 1339.735 cM for 'Kiyomi' and 'Murcott', respectively. Phenotyping for MS and polyembryony was performed. The genotype-trait association study identified a genomic region on LG8 which was significantly associated with MS, and a genomic region on LG1 which was significantly associated with polyembryony. Annotation of the identified region for MS revealed 19 candidate genes. One SNP KASPar marker was developed and fully validated for each trait.