Development of EST-SSR markers and association analysis of floral scent in tree peony

Tree peony (Paeonia Sect. Moutan) is widely grown as an ornamental plant and is known for its ‘National Beauty and Heavenly Fragrance’. However, the genetic improvement of floral scent traits is fundamentally limited by the paucity of genomic marker resources. Given the large-scale development of microsatellite markers using transcriptome sequences, we conducted an association analysis to identify microsatellite markers associated with main floral components. A total of 8,443 EST-SSRs were identified, with an average of one SSR per 15.55 kb. Di-nucleotide repeats were the most plentiful repeat type, followed by tri-, tetra-, penta- and hexa-nucleotide repeats. Among 969 synthesized primer pairs, 499 were successfully amplified and 136 showed polymorphism which exhibited relatively high levels of transferability were observed across 12 individuals in the genus Paeonia, thus expanding their utility. Subsequently, 31 polymorphic EST-SSR markers were selected for the analysis of genetic diversity and population structure, resulting in 176 alleles; the polymorphism information content (PIC) varied from 0.31 to 0.87. According to the population structure analysis, the association varieties were divided into three subgroups. Finally, single marker-trait association analysis based on mixed linear model (MLM) identified 29 significant associations, involving five floral components with 18 SSRs and a low proportion of phenotypic variance ranging from 4.57 to 23.46%. The newly developed markers were valuable resources that could provide a powerful genomic tool for genetic studies and accelerate the process of molecular marker-assisted breeding of tree peony floral scent.