Molecular Dissection of Quantitative Variation in Bermudagrass Hybrids (Cynodon dactylon x transvaalensis): Morphological Traits.

Bermudagrass (Cynodon (L.)) is the most important warm-season grass grown for forage or turf. It shows extensive variation in morphological characteristics and growth attributes, but the genetic basis of this variation is little understood. Detection and tagging of quantitative trait loci (QTL) affecting above-ground morphology with diagnostic DNA markers would provide a foundation for genetic and molecular breeding applications in bermudagrass. Here, we report early findings regarding genetic architecture of foliage (canopy height, HT), stolon (stolon internode length, ILEN and length of the longest stolon LLS), and leaf traits (leaf blade length, LLEN and leaf blade width, LW) in 110 F1 individuals derived from a cross between Cynodon dactylon (T89) and C. transvaalensis (T574). Separate and joint environment analyses were performed on trait data collected across two to five environments (locations, and/or years, or time), finding significant differences (P < 0.001) among the hybrid progeny for all traits. Analysis of marker-trait associations detected 74 QTL and 135 epistatic interactions. Composite interval mapping (CIM) and mixed-model CIM (MCIM) identified 32 main effect QTL (M-QTL) and 13 interacting QTL (int-QTL). Colocalization of QTL for plant morphology partially explained significant correlations among traits. M-QTL qILEN-3-2 (for ILEN; R2 = 11-19%), qLLS-7-1 (for LLS; R2 = 13-27%), qLEN-1-1 (for LLEN; R2 = 10-11%), and qLW-3-2 (for LW; R2 = 10-12%) were 'stable' across multiple environments, representing candidates for fine mapping and applied breeding applications. QTL correspondence between bermudagrass and divergent grass lineages suggests opportunities to accelerate progress by predictive breeding of bermudagrass.

Genotypic and phenotypic evaluation of off-type grasses in hybrid Bermudagrass [Cynodon dactylon (L.) Pers. x C. transvaalensis Burtt-Davy] putting greens using genotyping-by-sequencing and morphological characterization.

BACKGROUND: Interspecific hybrid bermudagrass [Cynodon dactylon (L.) Pers. x C. transvaalensis Burtt-Davy] is one of the most widely used grasses on golf courses, with cultivars derived from 'Tifgreen' or 'Tifdwarf' particularly used for putting greens. Many bermudagrass cultivars established for putting greens can be genetically unstable and lead to the occurrence of undesirable off-type grasses that vary in phenotype. The objective of this research was to genetically and phenotypically differentiate off-type grasses and hybrid cultivars. Beginning in 2013, off-type and desirable hybrid bermudagrass samples were collected from golf course putting greens in the southeastern United States and genetically and phenotypically characterized using genotyping-by-sequencing and morphology. RESULTS: Genotyping-by-sequencing determined that 11% (5) of off-type and desirable samples from putting greens were genetically divergent from standard cultivars such as Champion, MiniVerde, Tifdwarf, TifEagle, and Tifgreen. In addition, genotyping-by-sequencing was unable to genetically distinguish all standard cultivars from one another due to their similar origin and clonal propagation; however, over 90,000 potentially informative nucleotide variants were identified among the triploid hybrid cultivars. CONCLUSIONS: Although few genetic differences were found in this research, samples harvested from golf course putting greens had variable morphology and were clustered into three distinct phenotypic groups. The majority of off-type grasses in hybrid bermudagrass putting greens were genetically similar with variable morphological traits. Off-type grasses within golf course putting greens have the potential to compromise putting surface functionality and aesthetics.

SSR-enriched genetic linkage maps of bermudagrass (Cynodon dactylon × transvaalensis), and their comparison with allied plant genomes.

KEY MESSAGE: We report SSR-enriched genetic maps of bermudagrass that: (1) reveal partial residual polysomic inheritance in the tetraploid species, and (2) provide insights into the evolution of chloridoid genomes. This study describes genetic linkage maps of two bermudagrass species, Cynodon dactylon (T89) and Cynodon transvaalensis (T574), that integrate heterologous microsatellite markers from sugarcane into frameworks built with single-dose restriction fragments (SDRFs). A maximum likelihood approach was used to construct two separate parental maps from a population of 110 F1 progeny of a cross between the two parents. The T89 map is based on 291 loci on 34 cosegregating groups (CGs), with an average marker spacing of 12.5 cM. The T574 map is based on 125 loci on 14 CGs, with an average marker spacing of 10.7 cM. Six T89 and one T574 CG(s) deviated from disomic inheritance. Furthermore, marker segregation data and linkage phase analysis revealed partial residual polysomic inheritance in T89, suggesting that common bermudagrass is undergoing diploidization following whole genome duplication (WGD). Twenty-six T89 CGs were coalesced into 9 homo(eo)logous linkage groups (LGs), while 12 T574 CGs were assembled into 9 LGs, both putatively representing the basic chromosome complement (x = 9) of the species. Eight T89 and two T574 CGs remain unassigned. The marker composition of bermudagrass ancestral chromosomes was inferred by aligning T89 and T574 homologs, and used in comparisons to sorghum and rice genome sequences based on 108 and 91 significant blast hits, respectively. Two nested chromosome fusions (NCFs) shared by two other chloridoids (i.e., zoysiagrass and finger millet) and at least three independent translocation events were evident during chromosome number reduction from 14 in the polyploid common ancestor of Poaceae to 9 in Cynodon.

The genetic and phenotypic variability of interspecific hybrid bermudagrasses (Cynodon dactylon (L.) Pers. × C. transvaalensis Burtt-Davy) used on golf course putting greens.

MAIN CONCLUSION: Some interspecific hybrid bermudagrass cultivars used on golf course putting greens are genetically unstable, which has caused phenotypically different off-type grasses to occur in production nurseries and putting surfaces. Management practices to reduce the occurrence of off-type grasses in putting green surfaces and the effect they can have on putting quality and performance need to be researched until genetically stable cultivars are developed. Golf course putting green surfaces in subtropical and tropical climates are typically planted with an interspecific hybrid bermudagrass (Cynodon dactylon (L.) Pers. × C. transvaalensis Burtt-Davy), because of the superior putting quality and performance of these cultivars. 'Tifgreen' was one of the first interspecific hybrids developed for putting green use in lieu of common bermudagrass. However, off-type grasses began appearing in established Tifgreen stands soon after commercial release. Off-type grasses are those with different morphology and performance when compared to the surrounding, desirable cultivar. Off-types have the potential to decrease surface uniformity, which negatively affects putting surface quality. However, several unique off-types from Tifgreen have been selected as commercial cultivars, the first being 'Tifdwarf'; then 'Floradwarf', 'MS-Supreme', 'Pee Dee-102', and 'TL-2', identified later. The cultivars 'Champion Dwarf', 'P-18', 'RJT', and 'Emerald Dwarf' were subsequently selected as off-types in Tifdwarf. The naturally occurring off-types and cultivars that have been identified within the Tifgreen family have widely differing phenotypes; however, they are reported to be genetically similar, supporting the hypothesis that their occurrence is a result of somatic mutations. Genetic instability in currently available commercial cultivars is likely to lead to the continued presence of off-types in production nurseries and putting greens. Additional research is needed to understand the nature of genetic instability in Tifgreen-derived cultivars and how to manage its consequences to develop new cultivars, but also strategies for eradication of off-types in pedigree nursery production and end-site putting greens.

Synonymous mutation gene design to overexpress ACCase in creeping bentgrass to obtain resistance to ACCase-inhibiting herbicides.

Overexpression of a native gene can cause expression of both introduced and native genes to be silenced by posttranscriptional gene silencing (PTGS) mechanisms. PTGS mechanisms rely on sequence identity between the transgene and native genes; therefore, designing genes with mutations that do not cause amino acid changes, known as synonymous mutations, may avoid PTGS. For proof of concept, the sequence of acetyl-coA carboxylase (ACCase) from creeping bentgrass (Agrostis stolonifera L.) was altered with synonymous mutations. A native bentgrass ACCase was cloned and used as a template for the modified gene. Wild-type (WT) and modified genes were further modified with a non-synonymous mutation, coding for an isoleucine to leucine substitution at position 1781, known to confer resistance to ACCase-inhibiting herbicides. Five-hundred calli of creeping bentgrass 'Penn A-4' were inoculated with Agrobacterium containing either the WT or modified genes, with or without the herbicide-resistance mutation. Six herbicide-resistant-transgenic events containing the modified gene with the 1781 mutation were obtained. Transcription of the modified ACCase was confirmed in transgenic plants, showing that gene-silencing mechanisms were avoided. Transgenic plants were confirmed to be resistant to the ACCase-inhibiting herbicide, sethoxydim, providing evidence that the modified gene was functional. The result is a novel herbicide-resistance trait and shows that overexpression of a native enzyme with a gene designed with synonymous mutations is possible.