Abstract
Stylosanthes seabrana (Maass and ‘t Mannetje) (2n = 2x = 20), commonly known as Caatinga stylo, is an important tropical perennial forage legume. In nature, it largely co-exist with S. scabra, an allotetraploid (2n = 4x = 40) species, sharing a very high similarity for morphological traits like growth habit, perenniality, fruit shape and presence of small appendage at the base of the pod or loment. This makes the two species difficult to distinguish morphologically, leading to chances of contamination in respective germplasm collections. In present study, 10 S. seabrana accessions were discovered from the existing global germplasm stock of S. scabra represented by 48 diverse collections, utilizing sequence-tagged-sites (STS) genome-specific markers. All the newly identified S. seabrana accessions displayed STS phenotypes of typical diploid species. Earlier reports have conclusively indicated S. seabrana and S. viscosa as two diploid progenitors of allotetraploid S. scabra. With primer pairs SHST3F3/R3, all putative S. seabrana yielded single band of ~550 bp and S. viscosa of ~870 bp whereas both of these bands were observed in allotetraploid S. scabra. Since SHST3F3/R3 primer pairs are known to amplify single or no band with diploid and two bands with tetraploid species, the amplification patterns corroborated that all newly identified S. seabrana lines were diploid in nature. Flow cytometric measurement of DNA content of the species, along with distinguishing morphological traits such as flowering time and seedling vigour, which significantly differ from S. scabra, confirmed all identified lines as S. seabrana. These newly identified lines exhibited high level of similarity among themselves as revealed by RAPD and STS markers (>92% and 80% respectively). Along with the enrichment in genetic resources of Stylosanthes, these newly identified and characterized accessions of S. seabrana can be better exploited in breeding programs targeted to quality.
Similar content being viewed by others
References
Williams, R. J., Reid, R., Schultze-Karft, R., Sousa Costa, N., & Thomas, B. D. (1984). Natural distribution of Stylosanthes. In H. M. Stace & L. A. Edye (Eds.), The biology agronomy of Stylosanthes (pp. 73–101). Sydney, Australia: Academic Press.
Kirkbride, J. H., & de Kirkbride, C. G. (1985). Typification of Stylosanthes (Leguminosae) and its sections. Taxon, 36, 455–458.
Cameron, D. F. (1967). Chromosome number and morphology of some introduced Stylosanthes species. Australian Journal of Agricultural Research, 18, 375–379.
Edye, L. A. (1987). Potential of Stylosanthes for improving tropical grasslands. Outlook Agriculture, 16, 124–130.
Thomas, D. (1984). Global adventures in Stylosanthes. I. South America. In H. M. Stace & L. A. Edye (Eds.), The biology and agronomy of Stylosanthes (pp. 451–460). Sydney, Australia: Academic Press.
Cameron, D. F., Edye, L. A., Chakraborty, S., Manners, J. M., & Liu, C. J. (1996). An integrated program to improve anthracnose resistance in Stylosanthes—a review. Retrieved from http://www.regional.org.au/au/asa/1996/contributed/112cameron.htm.
Chandra, A., Pathak, P. S., & Bhatt, R. K. (2006). Stylo research in India: Prospects and challenges ahead. Current Science, 90, 915–921.
Ramesh, C. R., Bhag Mal, Hazara, C. R., Sukanya, D. H., Ramamurthy, V., & Chakraborty, S. (1997). Status of Stylosanthes development in other countries. III. Stylosanthes development and utilization in India. Tropical Grasslands, 31, 467–475.
Kazan, K., Manners, J. M., & Cameron, D. F. (1993). Genetic variation in agronomically important species of Stylosanthes determined using random amplified polymorphic DNA markers. Theoretical and Applied Genetics, 85, 882–888.
Curtis, M. D., Nourse, J. P., & Manners, J. M. (1995). Nucleotide sequence of a cationic peroxidase gene from the tropical forage legume Stylosanthes humilis. Plant Physiology, 108, 1303–1304.
Liu, C. J., & Musial, J. M. (1997). Stylosanthes sp. aff. S. scabra: a putative diploid progenitor of Stylosanthes scabra (Fabaceae). Plant Systematics and Evolution, 208, 99–105.
Ma, Z. Y., Chandra, A., Musial, J. M., & Liu, C. J. (2004). Molecular evidence that Stylosanthes angustifolia is the third putative diploid progenitor of the hexaploid S. erecta (Fabaceae). Plant Systematics and Evolution, 248, 171–176.
vander Stappen, J., Weltjens, I., Campenhout, S. V., & Volckaert, G. (1999). Genetic relationships among Stylosanthes species as revealed by sequence-tagged site markers. Theoretical and Applied Genetics, 98, 1054–1062.
Liu, C. J., Musial, J. M., & Thomas, B. D. (1999). Genetic relationships among Stylosanthes species revealed by RFLP and STS analyses. Theoretical and Applied Genetics, 99, 1179–1186.
Mannetje, L’t. (1984). Considerations on the taxonomy of the genus Stylosanthes. In H. M. Stace & L. A. Edye (Eds.), The biology and agronomy of Stylosanthes (pp. 1–21). Sydney, Australia: Academic Press.
Edye, L. A., & Hall, T. J. (1993). Development of new Stylosanthes cultivars for Australia from naturally occurring genotypes. In Proceedings of the XVIIth international grassland congress, Palmerstonpp North, New Zealand (pp. 2159–2161).
Liu, C. J. (1997). Geographical distribution of genetic variation in Stylosanthes scabra revealed by RAPD analysis. Euphytica, 98, 21–27.
Tewari, S., & Chandra, A. (2008). Genetical assessment of diploid progenitors of S. scabra by isozyme, RAPD and STS markers: A possible strategy for improvement of drought tolerant allo-tetraploid S. scabra species. Euphytica, 162, 39–50.
Chandra, A., Majumdar, A. B., Suresh, G., Bhatt, R. K., Roy, A. K., & Pathak, P. S. (2004). Evaluation of Stylosanthes germplasm: A multidisciplinary approach to identify lines for wider use in India. Paper in 4th international crop science congress, Brisbane, Australia.
Hall, T. J., & Glatzle, A. (2004). Cattle production from Stylosanthes pastures. In S. Chakraborty (Ed.), High yielding anthracnose—Resistant Stylosanthes for agricultural systems (pp. 51–64). Canberra, Australia: ACIAR.
Phaikaew, C., Ramesh, C. R., Kexian, Y., & Stur, W. (2004). Utilization of Stylosanthes as a forage crop in Asia. In S. Chakraborty (Ed.), High yielding anthracnose—Resistant Stylosanthes for agricultural systems (pp. 153–158). Canberra, Australia: ACIAR.
Matzk, F., Meister, A., & Schubert, I. (2000). An efficient screen for reproductive pathways using mature seeds of monocots and dicots. Plant Journal, 21, 97–108.
Arumuganathan, K., & Earle, E. D. (1991). Estimation of nuclear DNA content of plants by flow cytometry. Plant Molecular Biology Reporter, 9, 229–241.
Liu, C. J., & Musial, J. M. (1995). Restriction fragment length polymorphism detected by cDNA and genomic DNA clones in Stylosanthes. Theoretical and Applied Genetics, 91, 1210–1213.
Liu, C. J., Musial, J. M., & Smith, F. W. (1996). Evidence for a low level of genomic specificity of sequence tagged sites in Stylosanthes. Theoretical and Applied Genetics, 93, 864–868.
Manners, J. M., McIntyre, C. L., & Nourse, J. P. (1995). Cloning and sequence of a cDNA encoding phenylalanine ammonia-lyase from the tropical forage legume Stylosanthes humilis. Plant Physiology, 108, 1301–1302.
Reddy, A. S., Ranganathan, B., Haisler, R. M., & Swize, M. A. (1996). A cDNA encoding acyl-CoA-binding protein from cotton. Plant Physiology, 111, 348.
Smith, F. W., Ealing, P. M., Hawkesford, M. J., & Clarkson, D. T. (1995). Plant members of a family of sulfate transporters reveal functional subtypes. Proceedings of the National Academy of Sciences of the United States of America, 92, 9373–9377.
Rohlf, F. J. (1998). NTSYS-pc: Numerical taxonomy and multivariate analysis system, in Version 2.1. New York: Exeter Software, Applied Biostatistics Setauket.
Yap, I. V., & Nelson, R. J. (1996). Winboot: A program for performing bootstrap analysis of binary data to determine the confidence limits of UPGMA-based dendrograms. Philippines: IRRI.
Kaushal, P., Agrawal, A., Malaviya, D. R., Siddiqui, S. A., & Roy, A. K. (2009). Ploidy manipulation in guinea grass (Panicum maximum Jacq., Poaceae) utilizing a hybridization-supplemented apomixis-components partitioning approach (HAPA). Plant Breeding (in press).
Dolezel, J., & Bartos, J. (2005). Plant DNA flow cytometry and estimation of nuclear genome size. Annals of Botany, 95, 99–110.
Tuna, M., Vogel, K. P., Arumuganathan, K., & Gill, K. S. (2001). DNA content and ploidy determination of bromegrass germplasm accessions by flow cytometry. Crop Science, 41, 1629–1634.
Kaushal, P., Roy, A. K., Khare, A., Malaviya, D. R., Zadoo, S. N., & Choubey, R. N. (2007). Crossability and characterization of interspecific hybrids between sexual Pennisetum glaucum (pearl millet) and a new cytotype (2n = 56) of apomictic P. squamulatum. Cytologia, 72, 111–118.
Talent, N., & Dickinson, T. A. (2005). Polyploidy in Crataegus and Mespilus (Rosaceae, Maloideae): Evolutionary inferences from flow cytometry of nuclear DNA amount. Canadian Journal of Botany, 83, 1268–1304.
Date, R. A., Edye, L. A., & Liu, C. J. (1996). Stylosanthes sp. aff. scabra—a potential new forage plant for northern Australia. Tropical Grasslands, 30, 133.
Chandra, A., Bhatt, R. K., Majumdar, A. B., Kumar, S., Nagaich, D., & Kumar, K. (2008). Identification, evaluation and molecular characterization of Stylosanthes seabrana—A potential and nutritious range legume having wider applicability in India. In Proceedings of the XXI international Grassland congress and VIII international Rangeland congress, Huhhot, PR China, p. 363.
Acknowledgements
Authors are thankful to Director, IGFRI and Head, Crop Improvement Division for providing the necessary facilities to carry out the work. The seed materials received under ACIAR-stylo project and ILRI, Ethiopia through NBPGR are duly acknowledged. Financial support received to carry out the part of the work reported from ACIAR, Australia under the project (CS1/95/129) is duly acknowledged.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Chandra, A., Kaushal, P. Identification of Diploid Stylosanthes seabrana Accessions from Existing Germplasm of S. scabra Utilizing Genome-Specific STS Markers and Flow Cytometry, and Their Molecular Characterization. Mol Biotechnol 42, 282–291 (2009). https://doi.org/10.1007/s12033-009-9154-z
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12033-009-9154-z