Development of the Simple Sequence Repeat Marker PruG4RS for the Differentiation of Cherry Rootstocks

The use of clonally propagated Prunus sp. rootstocks in sweet cherry production is increasing as these rootstocks provide reduced tree size and precocity.  Numerous clonally propagated cherry rootstocks are in commercial production, such as the GiSelA series, or under test, such as the Michigan State University (MSU) series. Many of these dwarfing cherry rootstocks are difficult to differentiate based on morphology alone, therefore DNA markers that differentiate rootstocks are an important tool to verify identity and prevent mix-ups among these rootstocks during the vegetative propagation stage. The simple sequence repeat (SSR) marker PceGA59 was determined to uniquely distinguish the commercially available GiSelA rootstocks; however it did not distinguish all five of the MSU rootstock selections. In an attempt to meet this need a new SSR marker, PruG4RS, was developed based on an analysis of genome-wide genetic data for these rootstocks obtained using the RosBREED (www.rosbreed.org) Illumina Infinium® cherry SNP array. Two SNP markers were identified which were unique for each rootstock and sequence from the peach genome v1.0, available at the Genome Database for Rosaceae (www.rosaceae.org), was used to search for SSRs within the region surrounding the chosen SNPs. After identifying several potential SSR markers, those with the most specificity to the sequence of interest and least colinearity with additional scaffolds were selected and primer pairs designed for each. The most informative and highest quality SSR marker, PruG4RS, was developed from peach sequence upstream of the SNP marker ss490553174 on linkage group 4. The development of PruG4RS and its combined use with PceGA59 has successfully circumvented the limitations of each individual marker and proven effective for use as a “quality control” DNA diagnostic tool for the commercial GiSelA rootstocks as well as the MSU breeding program rootstock selections. RosBREED is funded by the USDA-NIFA-SCRI Award number 2014-51181-22378.