Golf Course Management

JUN 2018

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70 GOLF COURSE MANAGEMENT 06.18 phenotypic variability that can occur in an individual putting green, from golf course-to- golf course as well as cultivar-to-cultivar. Dif - ferences in internode and leaf length within the same putting surface can lead to decreased turfgrass density and reductions in putting surface quality and playability (6). Genetic characterization e inconsistencies of morphological mea- surements among hybrid bermudagrasses used on putting greens suggest genetic techniques may be more accurate in evaluating the di - versity of these grasses. Several methods have been used to explore the genotypic differences among bermudagrass cultivars and off-type grasses. e methodology used in these previ - ous experiments has been detailed elsewhere (6), but the majority of experiments share the same conclusion: Cultivars within the Tifgreen- derived family are not readily distinguished from one another using molecular marker technology (that is, DAF, AFLP and SSR). One molecular marker method, geno - typing-by-sequencing, had not been used to identify cultivars and off-type grasses. Geno - Multidimensional scaling plots were then generated from these variants to illustrate the variation among samples (Figure 4). e genotyping-by-sequencing results were surprising because the majority of sam - ples harvested from golf courses clustered with the standard cultivars Champion, MiniVerde, Tifdwarf, TifEagle and Tifgreen in the mul - tidimensional scaling plot (Figure 4). is clustering suggested that these samples were genetically similar to those cultivars. Only five (~11%) of the 47 unknown samples were genetically divergent from the standard cul - tivars. e ultradwarf cultivars were also ge- netically similar to Tifgreen and Tifdwarf, but genotyping-by-sequencing separated Tifway from the other hybrid cultivars. Our results using genotyping-by-sequencing were similar to previous molecular genetics research, which also failed to readily distinguish among ultra - dwarf cultivars and most off-type grasses. So why did the majority of grasses included in our experiment (and others) exhibit variable morphological characteristics while being ge - netically similar? is is still a "known un- known" with the off-type issue. However, the majority of bermudagrass putting green cul - tivars were selected from other bermudagrass cultivars (6), and the off-type grasses studied here were also selected from existing plant - ings. e difference in morphology could be driven by differential gene expression influ - enced by environment or management prac- tices (2,8). e intense management practices implemented on ultradwarf putting greens could result in the up- or down-regulation of genes that control important turfgrass char - acteristics (that is, internode and leaf length). However, no research has been conducted to explore this hypothesis. Off-type responses to nitrogen and trinexapac-ethyl We evaluated the response of off-type grasses and ultradwarf cultivars to nitrogen and trinexapac-ethyl applications. ree ul - tradwarf cultivars (Champion, MiniVerde and TifEagle) and three off-type grasses were treated with increasing nitrogen rates (0 to 1 pound nitrogen 1,000 square feet/week). One off-type was selected from each distinct morphological cluster. e grasses were estab - lished in greenhouse culture in a sand/peat mix meeting USGA root-zone recommenda - tions. Daily clipping was suspended at the time of initial nitrogen treatment application Figure 3. Photographs of samples representative of each morphological cluster. Grasses in cluster 1 had significantly longer internode lengths than those in clusters 2 and 3. Grasses in cluster 3 had significantly longer leaves than those in clusters 1 and 2. Photos by Eric Reasor typing-by-sequencing is capable of efficiently identifying large numbers of single nucleotide (DNA base) variants (1). In addition, genotyp - ing-by-sequencing has been used with success in switchgrass (3), wheat and barley (5). Based on its robustness and its successful use in other grasses, we hypothesized that genotyping-by- sequencing might be able to identify genetic variation among off-type grasses and hybrid bermudagrasses used on putting greens. DNA was isolated from bermudagrass samples and then sent to the Cornell Uni - versity Institute for Biotechnology in Ithaca, N.Y., for genotyping-by-sequencing analysis. Samples of known Champion, MiniVerde, Tifdwarf, TifEagle, Tifgreen and Tifway were included as standards, along with 47 off-type and desirable grasses from putting greens. e initial step in GBS involved a restriction en - zyme "cutting" the DNA into small pieces. ose small pieces of DNA were then fitted with barcode adapters for sequencing and sample recognition. After the small pieces of DNA were sequenced, the bioinformatics analysis pipeline sorted, indexed and merged the reads to determine nucleotide variants.

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