Application of SSRs Developed and Characterized for Buffalograss to Assess Sequence Variation.

Buffalograss [Buchloë dactyloides (Nutt.) Engel. syn Bouteloua dactyloides (Nutt.) Columbus] is a low input, sustainable turfgrass species. Until recently, large numbers of robust and informative markers were unavailable to assess genetic variation in economically important species. Next–generation sequencing (NGS) technologies were used to develop simple sequence repeat (SSR) markers at reduced cost and effort of traditional methods. The transcriptomes of buffalograss cultivars Prestige and 378 were sequenced. A total of 79.6 Mb and 9.5 Gb of transcriptome sequence data was generated from 454 and Illumina sequencing, respectively. A hybrid de novo assembly strategy was used to construct longer continuous sequences of Prestige. The Prestige transcriptome was mined for SSR containing sequences by the software program RepeatMasker. An in silico approach was used to identify SSRs that differed by copy number between 378 and Prestige. A total of 96 SSR primer pairs were designed. One application of these SSRs was to investigate the origin of an increased percentage of males occurring in subsequent growing seasons in female rows of an experimental breeder field. For enhanced aesthetics, cultural practices, and increased seed yield, female types are preferred and as such it would benefit the turfgrass industry to reduce the male to female plant ratio. For this study, a subset of 24 SSR primer pairs was screened and two primer pairs (B4 and E7) were selected for their ability to differentiate the one male parent from three female parents in the field where two rows of the male were planted in between eight rows of females. Leaf tissue from male types within the female rows and both field-collected and greenhouse-grown parental tissue were analyzed. Buffalograss is primarily a dioecious plant. The majority of the invasive male types had the same SSR profile as the female parents indicating that they were likely the result of monoecy that can result in male structures on plants that were previously female. Some male types had the male parent SSR profile that suggests aggressive stolon growth into the female rows. Additionally, a small percentage of male types had ‘alleles’ in common with both male and female parents indicating hybrid crosses. The appearance of ‘alleles’ not found in either the male or female parents suggests pollen contribution from outside the production field.