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United States Department of Agriculture

Agricultural Research Service

Research Project: RESEARCH, ACQUISITION, MANAGEMENT, AND DOCUMENTATION OF PLANT GENETIC RESOURCES Title: Using molecular markers to analyze genetic diversity in forage, turf, and biofuel crops

item Kisha, Theodore

Submitted to: International Symposium on Forage, Turfgrass and Biofuel Germplasm Research
Publication Type: Proceedings
Publication Acceptance Date: June 30, 2010
Publication Date: October 9, 2010
Citation: Kisha, T.J. 2010. Using molecular markers to analyze genetic diversity in forage, turf, and biofuel crops. International Symposium on Forage, Turfgrass and Biofuel Germplasm Research. October 9-13, 2010 Yangling City, China.

Interpretive Summary: The presentation reports on molecular diversity analyses on forage legumes and grasses, turfgrass species, and on safflower, an important potential biofuel crop. This work has helped to recognize areas for in situ conservation of genetic resources and has provided evidence for the classification of an endangered species and supports efforts to restore grazingland ecosystems and develop biofuels - critical challenges to both US and China.

Technical Abstract: The ex situ conservation of plant genetic resources has received worldwide attention for many years. Agriculture in the USA, as well as in many other countries, is primarily based on crop plants native to other parts of the world. Improvement of major crops for yield, quality, and resistance to biotic and abiotic stresses must rely on new genes found in old or wild germplasm. Additionally, greater sensitivity to environmental issues has increased farmers’ dependence on host plant resistances as opposed to the use of pesticides to fight diseases and pest damage. Despite some useful breakthroughs in biotechnology allowing the tapping of tertiary gene pools for genes with specific purposes, primary and secondary gene pools are still the most important sources of genetic variation for plant breeders. Germplasm collections worldwide provide genes for today’s breeding efforts, while preserving other genes for future needs. Availability of genetic diversity is of limited use, however, without the identification and characterization of that diversity, so it can be exploited and applied in an efficient manner. The Western Regional Plant Introduction Station (WRPIS), part of the USDA–ARS National Plant Germplasm System (NPGS) maintains seed and clonal germplasm. Currently, WRPIS maintains over 85 000 accessions from over 4 042 plant species from 376 genera. The Germplasm Resources Information Network of the NPGS (GRIN, documents 33 repositories in the United States, comprising 536 426 accessions from 222 families, 2 201 genera, and 13 451 species (as of 2010), and these numbers increase daily. The System-wide Information Network for Genetic Resources (SINGER, lists collections maintaining almost 700 000 accessions from 12 of 15 international agricultural research centers of the Consultative Group on International Agricultural Research (CGIAR). Knowledgeable and effective exploitation of such large collections is difficult and their great size is a barrier to their use. Molecular markers have become an accepted and widely used tool for the measurement of genetic diversity. Molecular marker technology can be used to characterize the extent of diversity within a collection and for the development of collection management strategies, which may include the establishment of core collections, identification of redundancies or contamination, guidance for future collection efforts, and identification of gaps of ancestral crop relatives. In addition, analysis of worldwide genetic diversity can identify areas suited for the establishment of in situ conservation sites. A search of the AGRICOLA database ( by year using search terms ‘diversity’ and each respective molecular marker type shows that the amount of research dedicated to the analysis of genetic diversity using molecular markers grows each year. The advent of new marker technologies is soon followed by its application to diversity analysis, but old technology continues to be exploited. At the WRPIS, we have used various types of molecular markers to examine the diversity among accessions of forage legumes, forage and turf grasses, and safflower, a potential biofuel crop. These analyses have been on apomictic and selfing accessions, as well as obligate outcrossers, each presenting a unique perspective on analysis. Molecular studies of alfalfa have necessarily required population analysis. Using AFLP, RAPD, Microsatellite, and chloroplast markers from three populations of alfalfa (Medicago sativa L.), we showed that all four marker types revealed the same relative relationships among the three populations tested. AFLP markers provided information on the largest number of loci at the least cost. We have also used AFLP and microsatellite data to identify gene flow in wild populations of alfalfa and identified areas to be set aside for in situ conservation (Greene et al., 2008). Our results indicate that 40 – 50 plants are needed to provide the least variance for population comparison, depending on the closeness of the relationship between populations and the molecular marker used. However, we were able to discern population structure among cultivars and wild relatives with only 16 plants in each population using the software STRUCTURE (Pritchard et al., 2000). Transgressive segregation for quantitative traits, such as yield, in crop plants relies on the recombination of many different genes positively affecting that trait. Given the potential number of genetically distinct progeny from a single cross and the number of parents available for crossing, knowledge of parental characteristics and their relationship to one another is imperative. Kisha et al. (1997) showed that regression analysis of molecular genetic distance and mid-parent yield could predict the highest yielding progeny from crosses of soybean. Johnson et al. (2007) used AFLP markers to characterize 96 accessions from the USDA safflower collection representing seven world regions (the Americas, China, East Africa, East Europe, the Mediterranean, South Central Asia, and Southwest Asia). Regions differed in all pair-wise comparisons using a bootstrap procedure comparing distances within and among populations. There was a weak but significant correlation of the AFLP matrix with a phenotypic data matrix with 16 attributes consisting of oil, meal, and growth characteristics (r = 0.12, P = 0.05). This weak correspondence between molecular and phenotypic data underscores the need for both types of characterization to enhance management and utilization of germplasm. Analysis of the data with the software STRUCTURE and excluding the American accessions placed the 80 remaining accessions into eight likely groups. Afghanistan accessions formed a unique group, as did China, and Ethiopia. While there was some mixture, which would be inevitable given the amount of germplasm exchange which must have taken place in the past, the eight groups could be relatively distinguished as 1) Middle East, 2) Egypt/Sudan, 3) Ethiopia, 4) Afghanistan, 5) Europe, 6) India, 7) Pakistan/Iran, and 8) China. Crossing with parents chosen from among these groups can increase the potential for transgressive segregation and the production of unique, genetically distinct progeny. Bibliography Greene, S. L., Kisha, T. J., and N. Dzyubenko. 2008. Conserving Alfalfa Wild Relatives: Is Past Introgression with Russian Varieties Evident Today? Crop Sci. 48: 1853-1864. Johnson, R. C., Kisha, T. J., and M. A. Evans. 2007. Characterizing safflower germplasm with AFLP molecular markers. Crop Sci. 47: 1728-1736. Kisha, T. J., Sneller, C. H., and B. W. Diers. 1997. Relationship between genetic distance among parents and genetic variance in populations of soybean. Crop Sci. 37:1317-1325. Pritchard, J.K., Stephens, M, and P. Donnelly. 2000. Inference of population structure using multilocus genotype data. Genetics 155:945–959.

Last Modified: 11/27/2015
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