MSU Professor Developing Hybrid Superturf
EAST LANSING, Mich. -- In the future, people who care for and enjoy using golf courses, sports fields and parks may be able to worry less about how cold weather and drought affect the grass at their favorite recreational areas. With the development of new turfgrass hybrids by Suleiman Bughrara, professor in the Michigan State University (MSU) Department of Crop and Soil Sciences, the turfgrass industry may grow greener and stronger than ever before.
Since beginning his work at MSU in 1999, Bughrara has blazed new trails. Or, sometimes, frozen them. Bughrara completed a comprehensive snow mold study of more than 4,000 cloned varieties of creeping bentgrass by simulating winter for each plant. Twenty bentgrass varieties showed significant resistance to snow mold, one of the most detrimental diseases challenging the turfgrass industry. A follow-up study found six of the 20 snow-mold-resistant clones also showed resistance to dollar spot, the other main turf-troubling disease.
“Bentgrass has all the right characteristics of great turf but shows susceptibility to dollar spot and snow mold,” Bughrara said. “We will continue our work to examine ways of crossbreeding aesthetically pleasing varieties, such as colonial bentgrass, to maximize disease resistance.”
Bughrara and his research team continue making discovery after exciting discovery in turfgrass breeding. His work also includes ryegrass and fescue. Working to unlock the mystery of drought tolerance, Bughrara is integrating Atlas fescue genes (from semiarid regions of Morocco) into the perennial ryegrass genome. The hybrids have shown high drought tolerance in greenhouse research. Field evaluations and molecular mapping are under way.
“This is exciting work,” Bughrara said. “We are the only university in the United States doing this type of genetic work to improve cold and drought tolerance and disease resistance in turfgrass breeding.”
Bughrara sees potential breakthroughs in how all plants are grown, especially food plants.
“With the right location on a gene, we can create hybrids for cold and drought tolerance in other crops as well. Wheat, corn and rice that need less water to thrive? It could change the entire landscape of our food systems,” Bughrara said.
Bughrara’s position and several of his research projects are funded by Project GREEEN (Generating Research and Extension to meet Economic and Environmental Needs), Michigan’s plant agriculture initiative housed at MSU.
Founded in 1997, Project GREEEN is a cooperative effort between plant-based commodities and businesses together with the Michigan Agricultural Experiment Station, MSU Extension and the Michigan Department of Agriculture to advance Michigan’s economy through its plant-based agriculture. Its mission is to develop research and educational programs in response to industry needs, ensure and improve food safety, and protect and preserve the quality of the environment.
To learn more about Michigan’s plant agriculture initiative at MSU, visit www.greeen.msu.edu.
Contact: Natalie Ebig Scott
517-432-1555, ext. 157
Transferring gray snow mold and dollar spot resistance from colonial bentgrass to creeping bentgrass by inter-specific hybridization research proposal - 2006
Summary
Creeping bentgrass (Agrostis stolonifera) is susceptible to a wide range of diseases, such as gray snow mold caused by Typhula incarnata (Wu and Hsiang 1998), and dollar spot caused by Sclerotinia homoeocarpa (Murphy et al. 2000). There are no commercial cultivars found to be resistant to these diseases. Studies indicated that certain colonial bentgrass (Agrostis cappillaris) accessions are recognized as having good resistance to gray snow mold (Vergara and Bughrara 2005) and dollar spot (Belanger et al. 2004). Utilization of resistance from colonial bentgrass could be an efficient and economic method to improve disease resistance of creeping bentgrass. Our objective is to transfer disease resistance from colonial bentgrass to creeping bentgrass. The inter-specific hybrids between creeping and colonial bentgrass were obtained by artificial crossing. The backcross by using a creeping bentgrass cultivar as the recurrent parent will be undertaken for combining disease resistance and elite morphological characteristics in the progeny. Inter-pollination will be conducted in the resistant progeny to pyramid resistance genes against gray snow mold and dollar spot pathogens. Cytogenetic studies and molecular markers can be used in monitoring the introgression of the alien chromosome fragments in the resistant progeny, tagging the genes and making DNA markers for assist the breeding and selection.
Introduction
Creeping bentgrass is a cool-season grass species and commonly used in golf course, putting green, tees and fair ways due to its fine texture and adaptation to low mowing. However, creeping bentgrass is susceptible to a wide range of fungal diseases such as dollar spot and gray snow mold. Dollar spot is a foliar disease favored by high humidity, warm days, and cool nights. Gray snow mold is common in cold, humid conditions, especially under snow cover. These diseases are particularly important in northern regions of the United States where they cause considerable turf loss. Every year, at least one hundred million dollars worth of fungicides are applied to control these diseases, much of it for snow mold and dollar spot (Watson et al. 1992).
Certain colonial bentgrass accessions are widely recognized as having good resistance to gray snow mold (Vergara and Bughrara 2005) and dollar spot (Belanger et al. 2004). Transferring resistance from colonial to creeping bentgrass is a promising goal in bentgrass breeding programs. Hybridization between creeping and colonial bentgrass species can occur naturally in the field (Jones 1956) or in the greenhouse (Belanger et al. 2003), which makes it possible to utilize the colonial bentgrass disease resistance by genome introgression into the creeping bentgrass species. Assessment of genetic diversity among accessions of colonial bentgrass species could contribute to the elimination of undesirable duplications in the germplasm collection and increase the efficiency of research efforts. Researchers could select potential diverse resistant genes from different colonial bentgrass sources, and incorporate and pyramid them into creeping or other bentgrass species to enhance levels and durability of disease resistance. Utilization of the specific genes from related species was successfully practiced in many crops. Usually, the F1 hybrids were obtained by inter-specific cross, and then the backcrosses were made according to different goals to narrow down the undesired alien chromosome fragments, which can be detected by cytogenetic and molecular techniques.
Materials and methods
Plant material
A colonial bentgrass accession, resistant to gray snow mold and dollar spot, was used as one of the parents. The other parent is a creeping bentgrass cultivar (Fig.1). The F1 seeds which collected from the creeping bentgrass cultivar were planted in the greenhouse.
DNA extraction
The fresh leaves (about 3g) from F1 progeny were ground in liquid nitrogen and then the fine powder was transferred into 50 ml polypropylene tube. The extraction buffer (0.1M Tris Ph=8.0, 0.05MDTA pH=8.0, 0.5MNaCl, 1.24%SDS) was added with volume 1:1 to tissues powder. After incubating 65°C about 1 hour, the samples were purified by 1/3 total volume chloroform/isoamy (24:1), and then centrifuged at 2000rpm for 25 min. The upper phase was added into pre-cold pure ethanol. Wash DNA twice in 70% ethanol. Dilute the DNA in TE buffer after removing excess ethanol. Add RNAase and quantity the DNA.
DNA amplification and analysis
The SCAR marker was applied the creeping and colonial genome amplification. PCR products were run on a 1.5% (w/v) agarose gel and stained in ethidium bromide. Presence and absence of the SCAR band was visually scored and compared with samples of their parents. The AFLP protocol applied in this research was as described by Vos et al. (1995).
Cytological analysis
The inflorescences were taken from hybrids and fixed in the Farmer’s solution (3 ethanol: 1 glacial acetic acid). The inflorescences were fixed at room temperature overnight and stored at -20°C after replenishing the fixative. The pollen fertilities of hybrids were examined by aceto-carmine stainability.
Results
F1 hybrids of colonial and creeping bentgrass
The F1 hybrid seeds collected from creeping bentgrass were planted in the greenhouse for confirming the self-crossing seeds because it has been reported that bentgrass may have over 5% self-crossing rate (Fig.2.). The SCAR marker which can specific amplify colonial bentgrass genome was applied. After testing, 4 out of 20 seedlings are self-crossing seedling because no band from the colonial bentgrass genome can be specific amplified (Fig.3.). The other 16 plants were planted in the greenhouse for further backcross.
Meiosis observation and pollen fertility of hybrid plants
The hybrids pollen mother cells (PMCs) and pollen fertilities were studied. The hybrids (2n=28) were confirmed by chromosome pairing at meiosis observation. Chromosome pairing at metaphase I was typified by the mean values of 6.7 univalents, 8.1 bivalents, 1.1 trivalents, and 0.45 quadrivalent per PMC. Seven bivalents were present at the first metaphase in most of the cells tested (Fig.4), indicating that A2A2 genome of creeping and colonial bentgrass was highly homologous; A1A1 and A3A3 genomes of the both species were partially homologous. Pollen stainability, as a measure of pollen viability or fertility, was estimated for F1 progeny. The average pollen viability is 19.6% (Fig.5)
The results open possibility of introgression between these two species by backcrossing.
Expected results
Backcrossing
The progeny from the backcross will be obtained by using a creeping bentgrass cultivar as the recurrent parent. The BC1F1 progeny collected from creeping bentgrass will be subject to the molecular marker analysis to confirm the hybrids.
Screening disease resistance The BC1F1 progeny will be evaluated by artificial inoculation method. The snow mold disease screening will be performed in the cold room (4°C), and dollar spot disease screening will be conducted in the greenhouse. Inter-pollination will be conducted in the resistant progeny to pyramid resistance genes against gray snow mold and dollar spot pathogens
Analyzing genetic composition of BC1F1 using cytogenetic and molecular methods
FISH and GISH will be used in determination of the genetic composition of BC1F1. Meanwhile, molecular markers (AFLP, SSR and RAPD) will be applied for detecting alien chromosome segments in progeny genome.
References:
Belanger, F.C., S. Bonos, W.A. Meyer. 2004. Dollar Spot Resistant Hybrids between CreepingBentgrass and Colonial Bentgrass. Crop Sci. 44: 581-586.
Belanger, F.C., T.R. Meagher, P.R. Day, K. Plumley, W.A. Meyer. 2003. Interspecific hybridization between Agrostis stolonifera and related Agrostis species under field conditions. Crop Sci. 43: 240–246.
Murphy, J.A., W.A. Meyer, K.A. Plumley, B.B. Clarke, S.A. Bonos, W.K. Dickson, J.B.
Clark, D.A. Smith, T.J. Lawson. 2000. Performance of bentgrass cultivars and selections in New Jersey turf trials. In 1999 Rutgers Turfgrass Proceedings. New Jersey Turfgrass Expo, Atlantic City, NJ. vol. 31.
Vergara, G.V., S.S. Bughrara. 2005. Evaluation of Bentgrass for Resistance to Typhula incarnata Lasch. Plant Disease (submitted).
Vos, P., R. Hogers, M. Bleeker, M. Reijans, T.V.D. Lee, M. Hornes, A. Fritjers, J. Pot, J. Poleman, M. Kuiper, M. Zabeau.1995. AFLP: A new technique for DNA fingerprinting. Nucleic Acid Res. 23: 4407-4414.
Watson, J.R., H.E. Kaerwer, D.P. Martin.1992. The turfgrass industry. In D.V. Waddington et al. (ed.) Turfgrass. ASA, Madison, WI. pp. 29-88.
Wu, C. and T. Hsiang. 1998. Pathogenicity and Formulation of Typhula phacorrhiza, a Biocontrol Agent of Gray Snow Mold. Plant Dis. 82: 1003-1006.