Isolation, Characterization, and Evaluation of Microsatellite Loci for Cultivar Identification in the Ornamental Pampas Grass Cortaderia selloana

Abstract

We isolated and characterized microsatellite loci in the ornamental pampas grass Cortaderia selloana (Schult. & Schult. f.) Asch. & Graebn. for purposes of identifying cultivars and assessing genetic relationships among cultivars. Small insert genomic libraries were enriched for dinucleotide (CT)n and (CA)n repeats. Ninety clones were sequenced of which 76% contained at least one microsatellite with a basic motif greater than six repeat units. Nine primer pairs amplified 10 polymorphic and putatively disomic loci, and were used to genotype 88 individuals representing 17 named cultivars and four selections. In total, 93 alleles were detected with a maximum of two to 19 per locus. Effective number of alleles varied from 1.3 to 9.5. Observed heterozygosity ranged from 0.07 to 0.81. The 10 microsatellite loci distinguished the majority of pampas grass cultivars. An unweighted pair group method with arithmetic mean (UPGMA) cluster analysis, based on proportion of shared alleles among individuals, revealed groups of cultivars corresponding to origin and morphological characteristics. With few exceptions, individuals of a single cultivar clustered together with moderate to strong bootstrap support (greater than 50%). Interestingly, `Pumila' from Europe and the United States formed separate clusters indicating independent origins. A large, diverse cluster with low bootstrap support consisted of selections and cultivars sold as seed, rather than potted or bare-root clonal plants. Primers designed for C. selloana amplified microsatellite loci in other Cortaderia Stapf species concordant with phylogenetic relationships among the species. Cross-amplification was 100% in C. jubata (Lemoine ex Carrière) Stapf; 77% in C. pilosa (d'Urv.) Hack. and C. rudiuscula Stapf; 66% in C. fulvida (Buch.) Zotov; and 55% in C. richardii (Endl.) Zotov and C. toetoe Zotov.