Isolation and characterization of simple sequence repeat markers for the tetraploid forage grass Dactylis glomerata

Abstract

To isolate simple sequence repeat (SSR) markers from Dactylis glomerata, four SSR-enriched genomic libraries for different repeat motifs from genomic DNA of a Japanese orchardgrass variety, ÔAkimidori IIÕ, were constructed. A total of 969 unique SSR-containing clones were identified and used for primer design. A BLAST search revealed that 63 of the sequences showed significant homology with sequences of other plant species including 18 that showed significant gene homology. After screening of a panel consisting of eight orchardgrass individuals, 606 SSR markers showed polymorphism, 70 generated monomorphic bands and the other primers either generated multiple bands or did not provide any amplification product. In the 606 polymorphic SSR markers, most had a large number of alleles, ranging from 1 to 13 with an average of 4.5 alleles per SSR primer pair. The number of alleles per individual ranged from 1 to 4. These SSR markers developed in this study will have broad utility for genotyping applications in this species, including gene targeting, quantitative trait locus mapping, variety or species identification and marker-assisted selection.

Identification of orchardgrass (Dactylis glomerata L.) cultivars by using simple sequence repeat markers

“…Orchardgrass is an allogamous species that has a strong self-incompatibility system (Xie et al, 2012), which guarantees that alleles in each cycle of mating are reshuffled to cause a high degree of genetic variation within populations (Hirata et al, 2011). Therefore, it is difficult to identify orchardgrass cultivars based on physiological and morphological traits, because these traits are subjective and quantitative in practice, limited in number, and affected by environmental conditions and the experience of evaluators (Roldán-Ruiz et al, 2001;Bolaric et al, 2005).…”

“…It is auto-tetraploid, with a large genome (4312 Mbp) (Creber et al, 1994) and selfincompatibility system (Hirata et al, 2011). Compared to other forage material, the breeding of orchardgrass is hampered by its strong self-incompatibility system and severe inbreeding depression, which makes it difficult to produce inbred lines and construct fingerprinting (Xie et al, 2011).…”

“…Primers were screened from 88 SSR primer pairs, using a panel of 4 cultivars, i.e., Frode, Akaroa, Baoxing, and Amba. These primer sequences were kindly provided by Dr. Cai from Japan Grassland Farming and Forage Seed Association, Forage Crop Research Institute, Japan (Hirata et al, 2011 Table2. Orchardgrass SSR primers used in this study and the results detected in 32 orchardgrass cultivars.…”

Analysis of diversity and relationships among orchardgrass (Dactylis glomerata L.) accessions using start codon-targeted markers

“…In recent years, a number of molecular markers have been employed in orchardgrass for genetic diversity evaluation, genetic mapping, and quantitative trait locus analysis. These types of molecular techniques included random amplified polymorphic DNA (RAPD) (Kölliker et al, 1999;Tuna et al, 2004), amplified fragment length polymorphism (AFLP) (Reeves et al, 1998;Peng et al, 2008), inter-simple sequence repeat (ISSR) (Zeng et al, 2006), sequence-related amplified polymorphism (Zeng et al, 2008;Xie et al, 2011), simple sequence repeats (SSRs) (Hirata et al, 2011;Xie et al, 2011Xie et al, , 2012, and expressed sequence tag-SSRs (Xie et al, 2010;Bushman et al, 2011). These marker systems are useful for biodiversity analyses, phylogenetic studies, germplasm management, cultivar identification, and other applications (Semagn et al, 2006;Luo et al, 2010).…”

“…It has been widely used as forage on nearly every continent because of its high yield, good nutrition, adaptability, and shade tolerance (Jafari and Naseri, 2007;Sanada et al, 2007;Xie et al, 2010). Orchardgrass is one of the top 4 perennial forage grasses grown in North America, Oceania, Europe, and Asia (Lindner and Garcia, 1997;Gauthier et al, 1998;Hirata et al, 2011), and it plays an important role in the livestock industry and in the sustainable development of environments worldwide.…”