2012 Annual Report 1a.Objectives (from AD-416): To improve the existing sequencing and assembly of the Prunus domestica genome. 1b.Approach (from AD-416): DNA will be purified from Prunus domestica cultivars. Purified DNA will be outsourced to a sequencing service provider to obtain 3X genome coverage via long read sequencing (>500 base pair reads). 3.Progress Report: A more fully assembled genome will aid molecular marker development as well as genomic studies in this species. Prior to performing additional sequencing, computational analyses were performed using existing data. The results showed that the hexaploid nature of the genome was a limiting factor and that additional sequencing would not likely lead to significant improvements beyond the current plum-peach reference assembly. Thus, we moved to the next step of molecular marker development. A technician was hired to design a set of genome spanning molecular markers that should discriminate between the industry standard cultivar, 'Improved French', and an Eastern-type transgenic plum, 'HoneySweet'. Markers were designed against a set of Deletion/Insertion Polymorphisms and tested on approximately 70 plum cultivars. Twenty-four markers could effectively discriminate 'Improved French' from 'HoneySweet'. The data also revealed that 'Improved French' is distinctly different from the bulk of Prunus domestica germplasm. In addition, several DIP positions were found to have six distinct alleles representing each allele in the hexaploid genome. This is now being used to assess the precise genetic segregation pattern in Prunus domestica.