Molecular marker development and primary physical map construction for the tuber shape Ro gene locus in diploid potato (Solanum tuberosum L.)

Abstract

Potato (Solanum tuberosum L.) is an important crop worldwide that is consumed fresh and processed in many different ways. The tuber shape is a critical trait in potato breeding, because it directly affects processing quality. Previous studies have demonstrated that the Ro locus on chromosome X is the major locus controlling tuber shape, and the round shape allele, Ro, is dominant over the long shape allele, ro. However, no reports on the cloning or physical mapping of this locus have been published. In this study, whole genome sequencing and bacterial artificial chromosome (BAC) library construction were applied to develop molecular markers and construct a physical map of Ro. Eight round-tuber genotypes were used to construct the BAC library, which contains about 40,000 clones with an average insert size of 85 kb and represents approximately four haploid potato genomes. A total of 66 individuals with extreme round or long tubers were selected to construct a round-tuber shape DNA bulk and a long-tuber shape DNA bulk for WGS. Single-nucleotide polymorphisms (SNPs) identified from WGS data were used to develop nine polymorphic markers linked to Ro. These markers were used to screen the BAC library, and eight positive BAC clones were ultimately identified and then analyzed by restriction enzyme digestion and BAC end sequencing and used to develop markers and identify new positive BAC clones. Finally, a primary physical map of Ro was constructed using a chromosome walking strategy. This map provides an important reference for cloning the potato tuber shape gene and studying tuber shape variation.