Transferability of Genomic SSR Markers in Lens Species for Genus Characterization and Lentil (Lens culinaris Medik.) Breeding

Year 2025, Volume: 30 Issue: 1, 223 - 234, 23.06.2025

Brian Wakimwayi Koboyi , Merve Keklik , Özge Doğanay , Melike Bakır

https://doi.org/10.17557/tjfc.1623413

Abstract

Simple sequence repeats (SSRs) are pivotal for germplasm
characterization and crop improvement. However, lentil breeding faces
challenges due to the limited availability of markers. This study assessed
the transferability of genomic SSR markers developed in cultivated
lentils to wild Lens species. A total of 100 SSR loci were analyzed in 21
accessions representing all the Lens genus using PowerMarker software
v3.25, which identified 41 polymorphic loci. These loci generated 219
alleles, with an average of 5.34 per locus and a polymorphic information
content (PIC) value of 0.61. Notably, 52.2% of the polymorphic loci
were shared among all Lens species. Transferability rates varied across
species: 41% for Lens nigricans, 40% for Lens culinaris subsp.
tomentosus, 39% for Lens culinaris subsp. orientalis, 34% for Lens
culinaris subsp. odemensis, 34% for Lens ervoides, and 31% for Lens
lamottei. The identified polymorphic markers provide a valuable
resource for exploring the genetic diversity of wild lentil relatives and
the breeding of beneficial agronomic traits into new lentil cultivars.

Keywords

SSR , Lentil , Transferability , Genetic diversity , Crop improvement

Supporting Institution

Erciyes University Scientific Research Unit

Project Number

FCD-2021-10106

Thanks

We gratefully acknowledge Prof. Dr. Cengiz Toker and the Field Crops Department at Akdeniz University’s Faculty of Agriculture for supplying the plant material for this study. We also thank Betül-Ziya Eren Genome and Stem Cell Centre at Erciyes University for providing the laboratory facilities essential for the successful completion of this research.

References

• Ahmad, M., Fautrier, A. G., Burritt, D. J., & McNeil, D. L. (1997). Genetic diversity and relationships in Lens species and their F1 interspecific hybrids as determined by SDS‐PAGE, New Zealand Journal of Crop and Horticultural Science, 25(2), 99-108. https://doi.org/10.1080/01140671.1997.9513995
• Andeden, E. E., Baloch, F. S., Cakır, E., Toklu, F., & Ozkan, H. (2015). Development, characterization and mapping of microsatellite markers for lentil (Lens culinaris Medik.). Plant Breeding, 134(5), 589-598. https://doi.org/10.1111/pbr.12296
• Ates, D. (2019). Genetic diversity in lentil landraces revealed by diversity array technology (DArT). Turkish Journal of Field Crops, 24(2), 252-260. https://doi.org/10.17557/tjfc.656511
• Bakir, M. (2019). Transferability of newly developed genomic lentil SSR markers to Cicer species. Legume Research-An International Journal, 42(4), 479-484. https://doi.org/10.18805/LR-466
• Bakir, M., & Kahraman, A. 2019. Development of new SSR (simple sequence repeat) markers for lentils (Lens culinaris Medik.) from genomic library enriched with AG and AC microsatellites. Biochem Genet 57, 338–353. https://doi.org/10.1007/s10528- 018-9893-2
• Bakir, M., Demir, S., Yildirim, C., & Kahraman, A. (2024). Genomic SSR marker development in lentil (Lens culinaris Medik.) and assessment of cross-species/genera transferability to related legumes. Czech Journal of Genetics and Plant Breeding, 60(1). http://dx.doi.org/10.17221/57/2023-CJGPB
• Begna, T., & Yesuf, H. (2021). Genetic mapping in crop plants. Open J. Plant Sci. 6(1): 019-026. DOI: https://dx.doi.org/10.17352/ojps.000028
• Bett KE, Cook DR (2016) KnowPulse. (Accessed on July 01, 2021); Available online: https://knowpulse.usask.ca/ Botstein, D., White, R. L., Skolnick, M., & Davis, R. W. (1980). Construction of a genetic linkage map in man using restriction fragment length polymorphisms. Am. J. Hum. Genet. 32: 314-331
• Cevik, S., ÜNYAYAR, S., & ERGÜL, A. (2015). Genetic relationships between cultivars of Cicer arietinum and its progenitor grown in Turkey determined by using the SSR markers. Turkish Journal of Field Crops, 20(1). https://doi.org/10.17557/.87839
• Caracuta, V., Vardi, J., Paz, Y., & Boaretto, E. (2017). Farming legumes in the pre-pottery Neolithic: new discoveries from the site of Ahihud (Israel). PLoSOne 12:e0177859. https://doi.org/10.1371/journal.pone.0177859
• Choudhary, S., Sethy, N. K., Shokeen, B., & Bhatia, S. (2009). Development of chickpea EST-SSR markers and analysis of allelic variation across related species. Theoretical and Applied Genetics, 118, 591-608.de. https://doi.org/10.1007/s00122-008-0923- z
• Datta, S., Mahfooz, S., Singh, P., Choudhary, A. K., Singh, F., & Kumar, S. (2010). Cross-genera amplification of informative microsatellite markers from common bean and lentil for the assessment of genetic diversity in pigeonpea. Physiol.Mol. Biol. Plants 16: 123-134. https://doi.org/10.1007/s12298-010-0014-x
• Datta, S., Mahfooz, S., Sahil, P., Choudhary, A. K., Chaturvedi, S. K., & Nadarajan, N. (2013). Conservation of microsatellite regions across legume genera increases marker repertoire in pigeonpea. Australian Journal of Crop Science, 7(13), 1990-1997.
• Daudi, H., Shimelis, H., Mathew, I., Oteng‐Frimpong, R., Ojiewo, C., & Varshney, R. K. (2021). Genetic diversity and population structure of groundnut (Arachis hypogaea L.) accessions using phenotypic traits and SSR markers: implications for rust resistance breeding. Genetic Resources and Crop Evolution, 68, 581-604. https://doi.org/10.1007/s10722-020-01007-1
• de Oliveira, J. C., da Silva, A. L. D., & da Silva, L. M. et al. (2023). Novel Microsatellite Markers Derived from Arachis pintoi Transcriptome Sequencing for Cross-Species Transferability and Varietal Identification. Plant Mol Biol Rep. https://doi.org/10.1007/s11105-023-01402-9
• Demir, S., & Bakir, M. (2022). Development of new ssr markers by screening libraries enriched with AG and AC repeats with TG and TC repeats in lentil. European Journal of Science and Technology, (41), 54-58. https://doi.org/10.31590/ejosat.1075391
• Dhull, S. B., Kinabo, J., & Uebersax, M. A. (2022). Nutrient profile and effect of processing methods on the composition and functional properties of lentils (Lens culinaris Medik): a review. Legume Science 5, e156. https://doi.org/10.1002/leg3.156
• Dikshit, H. K., Singh, A., Singh, D., Aski, M. S., Prakash, P., Jain, N., ... & Sarker, A. (2015). Genetic diversity in Lens species revealed by EST and genomic simple sequence repeat analysis. PloS one, 10(9), e0138101. https://doi.org/10.1371/journal.pone.0138101
• Dissanayake, R., Braich, S., Cogan, N. O., Smith, K., & Kaur, S. (2020). Characterization of genetic and allelic diversity amongst cultivated and wild lentil accessions for germplasm enhancement. Frontiers in Genetics, 11, 546.
• FAOSTAT (2023) FAOSTAT. https://www.fao.org/faostat/en/#data/QCL. Accessed 18 June 2023 Felsenstein, J. (1993). PHYLIP (phylogeny inference package), version 3.5 c. Joseph Felsenstein.
• Ferguson, M. E., Robertson, L. D., Ford-Lloyd, B. V., Newbury, H. J., & Maxted, N. (1998). Contrasting genetic variation amongst lentil landraces from different geographical origins. Euphytica, 102, 265-273.
• Gupta, D., Taylor, P. W. J., Inder, P., Phan, H. T. T., Ellwood, S. R., Mathur, P. N., ... & Ford, R. (2012). Integration of EST-SSR markers of Medicago truncatula into intraspecific linkage map of lentil and identification of QTL conferring resistance to ascochyta blight at seedling and pod stages. Molecular breeding, 30, 429-439. https://doi.org/10.1007/s11032-011-9634-2
• Gutierrez-Gonzalez, J. J., García, P., Polanco, C., González, A. I., Vaquero, F., Vences, F. J., ... & Sáenz de Miera, L. E. (2022). Multi-species transcriptome assemblies of cultivated and wild lentils (Lens sp.) provide a first glimpse at the lentil pangenome. Agronomy, 12(7), 1619. https://doi.org/10.3390/agronomy12071619
• Hamwieh, A., Udupa, M., Sarker, A., Jung, C., & Baum, M. (2009). Development of new microsatellite markers and their application in the analysis of genetic diversity in lentils. Breed Sci. 59: 77–86.
• Hoffman, D. L., Muehlbauer, F. J., & Ladizinsky, G. (1988). Morphological variation in Lens (Leguminosae). Systematic Botany, 13(1), 87–96. https://doi.org/10.2307/2419244
• Jingade, P., Bhosale, L. V., Sanjayrao, J. A., Rajanna, R., Jain, M., & Ravikumar, R. L. (2014). Characterization of microsatellite markers, their transferability to orphan legumes and use in determination of genetic diversity among chickpea (Cicer arietinum L.) cultivars. Journal of Crop Science and Biotechnology, 17(3), 191-199.
• Koboyi, B. W., & Bakir, M. (2024). Integration of Novel SSR Markers into The Lentil (Lens culinaris Medik.) Genome. Turkish Journal of Field Crops, 29(1), 40-45. https://doi.org/10.17557/tjfc.1415823
• Konda, A. K., & Annapragada, H. (2024). Structural and functional genomic resources. In the Lentil Genome (pp. 195-216). Academic Press. https://doi.org/10.1016/B978-0-443-19409-2.00010-7
• Koul, P. M., Sharma, V., Rana, M., Chahota, R. K., Kumar, S., & Sharma, T. R. (2017). Analysis of genetic structure and interrelationships in lentil species using morphological and SSR markers. 3 Biotech 7:83. https://doi.org/10.1007/s13205-017- 0683-z
• Kumar, J., Srivastava, E., Singh, M., Kumar, S., Nadarajan, N., & Sarker, A. (2014). Diversification of indigenous gene-pool by using exotic germplasm in lentil (Lens culinaris Medikus subsp. culinaris). Physiol. Mol. Biol. Plants 20, 125–132. https://doi.org/10.1007/s12298-013-0214-2
• Lefort,F., Lally, M., Thompson, D., & Douglas, G.C. (1998). Morfolojical traits microsatellite fingerprinting and genetic relatedness of a stand of elite oaks (Q. Robur L.) At Tuallynally, Ireland. Silvae Genetica, 47, 5-6.
• Li, J. (2023). Draft genome assembly, organelle genome sequencing and diversity analysis of marama bean (Tylosema esculentum), the green gold of Africa (Doctoral dissertation, Case Western Reserve University).
• Liber, M., Oliveira, H. R., Duarte, I., & Maia, A. T. (2021). The history of lentil (Lens culinaris ssp. culinaris) domestication and spread as revealed by genotyping-bysequencing of wild and landrace accessions. Front. Plant Sci. 12. https://doi.org/10.3389/fpls.2021.628439
• Liu, K., & Muse, S. V. (2005). PowerMarker: an integrated analysis environment for genetic marker analysis. Bioinformatics 21:2128–2129. https://doi.org/10.1093/bioinformatics/bti282
• Mehandi, S., & Mishra, S. P. (2023). Genetic diversity and population structure analyses in mungbean (Vigna radiata L. Wilczek). J Food Chem Nanotechnol, 9(S1), S316-S322.
• Muehlbauer, F. J., & McPhee, K.E. (2005). Lentil (Lens culinaris Medik.). In: Singh R. J., Jauhar P. P., editors. Genetic Resources, Chromosome Engineering, and Crop Improvement: Grain Legumes. Taylor & Francis. pp. 268–280.
• Nei, M. (1973). The theory and estimation of genetic distance. Genetic Structure of Populations (ed. Morton, N. E.), pp. 45–54.
• Honolulu: University of Hawaii Press.Google ScholarPubMed Ningwal, R., Tripathi, M. K., Tiwari, S., Asati, R., Yadav, R. K., Tripathi, N., & Yasin, M. (2023). Identification of polymorphic SSR marker and diversity analysis in a set of desi chickpea genotypes. In Biological Forum-An International Journal (Vol. 15, No. 3, pp. 45-51).
• Odeny, D. A., Jayashree, B., Ferguson, M., Hoisington, D., Crouch, J., & Gebhardt, C. (2007). Development, characterization and utilization of microsatellite markers in pigeonpea. Plant Breeding, 126(2), 130-136. https://doi.org/10.1111/j.1439- 0523.2007.01324.x
• Ozkan, G., Haliloglu, K., Turkoglu, A., Ozturk, H. I., Elkoca, E., & Poczai, P. (2022). Determining genetic diversity and population structure of common bean (Phaseolus vulgaris L.) landraces from Türkiye using SSR markers. Genes, 13(8), 1410. https://doi.org/10.3390/genes13081410
• Peng, J. H., & Lapitan, N. L. (2005). Characterization of EST-derived microsatellites in the wheat genome and development of eSSR markers. Funct. Integr. Genomics 5: 80-96. https://doi.org/10.1007/s10142-004-0128-8
• Polanco, C., de Miera, L. E. S., González, A. I., García, P. G., Fratini, R., Vaquero, F., Vences, F. J., & De La Vega, M. P. (2019). Construction of a high-density interspecific (Lens culinaris × L. odemensis) genetic map based on functional markers for mapping morphological and agronomical traits, and QTLs affecting resistance to Ascochyta in lentil. PLoS ONE, 14, e0214409. https://doi.org/10.1371/journal.pone.0214409
• Rajendra, D., Ashutosh, S., Krishna Hari, D., & Madhav Prasad, P. (2019). Molecular characterization of high concentration of iron (Fe) and zinc (Zn) biofortified nepalese lentil (Lens culinaris Medikus Subspecies culinaris) accessions and their genetic diversity analysis through using simple sequence repeat (SSR) markers. BAOJ Biotech, 5, 041.
• Rajpal, V. R., Singh, A., Kathpalia, R., Thakur, R. K., Khan, M. K., Pandey, A., ... & Raina, S. N. (2023). The prospects of gene introgression from crop wild relatives into cultivated lentil for climate change mitigation. Frontiers in Plant Science, 14, 1127239. https://doi.org/10.3389/fpls.2023.1127239
• Reddy, M. R. K., Rathour, R., Kumar, N., Katoch, P., & Sharma, T. R. (2010). Cross‐genera legume SSR markers for analysis of genetic diversity in Lens species. Plant Breeding, 129(5), 514-518. https://doi.org/10.1111/j.1439-0523.2009.01723.x
• Renfrew, J. M. (1973). Palaeoethnobotany. Columbia Univ. Press, New York.
• Rubiales, D., & Fondevilla, S. (2012). Future prospects for Ascochyta blight resistance breeding in cool season food legumes. Frontiers in Plant Science, 3, 27. https://doi.org/10.3389/fpls.2012.00027
• Saidi, A., Sarvmeili, J., & Pouresmael, M. (2022). Genetic diversity study in lentil (Lens culinaris Medik.) Germplasm: a comparison of CAAT Box Derived Polymorphism (CBDP) and simple sequence repeat (SSR) markers. Biologia, 77(10), 2793- 2803. https://doi.org/10.1007/s11756-022-01089-5
• Saıdou, M., Wang, C., Alam, M. A., Chen, C., & Jı, W. (2016). Genetic analysis of powdery mildew resistance gene using SSR markers in common wheat originated from wild emmer (Triticum dicoccoides Thell). Turkish Journal of Field Crops, 21(1), 10-15. https://doi.org/10.17557/tjfc.83589
• Saxena, S., Kaila, T., Chaduvula, P. K., Singh, A., Singh, N. K., & Gaikwad, K. (2019). Novel chloroplast microsatellite markers in pigeonpea ('Cajanus cajan'L. Millsp.) and their transferability to wild'Cajanus' species. Australian Journal of Crop Science, 13(2), 185-191.
• Schuelke, M. (2000). An economic method for the fuorescent labelling of PCR fragments. Nat Biotechnol 18:233–234 Sharma S. K., Knox, M. X., & Ellis, T. H. N. (1996). AFLP analysis of the diversity and phylogeny of Lens and its comparison with RAPD analysis. Theor. Appl. Genet. 93: 751–758.
• Singh, A., Dikshit, H. K., Singh, D., Jain, N., Aski, M., Sarker, A., & Sharma, T. R. (2016). Use of expressed sequence tag microsatellite markers for exploring genetic diversity in lentil and related wild species. The Journal of Agricultural Science, 154(7), 1254-1269. https://doi.org/10.1017/S0021859615001252
• Singh, B., Malhotra, N., & Gupta, D. (2018). Widening the genetic base of cultivated gene pool following introgression from wild Lens taxa. Plant Breed. 137, 447–485. doi: 10.1111/pbr.12615
• Singh, D., Singh, C. K., Taunk, J., Jadon, V., Pal, M., & Gaikwad, K. (2019). Genome wide transcriptome analysis reveals vital role of heat responsive genes in regulatory mechanisms of lentil (Lens culinaris Medikus). Scientific reports, 9(1), 12976. https://doi.org/10.1038/s41598-019-49496-0
• Singh, M., Kumar, S., Basandrai, A. K., Basandrai, D., Malhotra, N., Saxena, D. R., Gupta, D., Sarker, A., & Singh, K. (2020). Evaluation and identification of wild lentil accessions for enhancing genetic gains of cultivated varieties. PLoS One, 15(3), e0229554. https://doi.org/10.1371/journal.pone.0229554
• Singh, M., Kumar, S., Mehra, R., Sood, S., Malhotra, N., Sinha, R., Jamwal, S., & Gupta, V. (2022). Evaluation and identification of advanced lentil interspecific derivatives resulted in the development of early maturing, high yielding, and disease-resistant cultivars under Indian agro-ecological conditions. Frontiers in Plant Science, 13, 936572. https://doi.org/10.3389/fpls.2022.936572
• Sharma, R., Chaudhary, L., & Kumar, M. (2023). Microsatellites based assessment of genetic diversity and population structure of indian lentil (Lens culinaris Medik.) genotypes. Biologia, 78(9), 2317-2328.
• Tamura K., Dudley J., Nei M., Kumar S. (2007): MEGA4:Molecular evolutionary genetics analysis (MEGA) software version 4.0. Molecular Biology and Evolution,24: 1596–1599. https://doi.org/10.1093/molbev/msm092
• Tantasawat, P., Trongchuen, J., Prajongjai, T., Jenweerawat, S., & Chaowiset, W. (2011). SSR analysis of soybean (Glycine max L. Merr.) genetic relationship and variety identification in Thailand. Australian J. Crop Sci. 5: 283-290
• Tiwari, K. K., Thakkar, N. J., Dharajiya, D.T. et al. (2021). Genome-wide microsatellites in amaranth: development, characterization, and cross-species transferability. 3 Biotech 11, 395. https://doi.org/10.1007/s13205-021-02930-5
• Topu, M., Sesiz, U., Bektas, H., Toklu, F., & Ozkan, H. (2023). Next-generation-sequencing-based simple sequence repeat (SSR) marker development and linkage mapping in lentil (Lens culinaris L.). Life, 13(7), 1579. https://doi.org/10.3390/life13071579
• Varshney, R. K., Graner, A., & Sorrells, M.E. (2005). Genic microsatellite markers in plants: features and applications. Trends Biotechnol. 23: 48-55. https://doi.org/10.1016/j.tibtech.2004.11.005
• Verma, P., Sharma, T., Srivastava, P., Abdin, M.Z., & Bhatia, S. (2014) Exploring genetic variability within lentil (Lens culinaris Medik.) and across related legumes using a newly developed set of microsatellite markers. Molecular Biology Reports: 1–19. https://doi.org/10.1007/s11033-014-3431-z
• Wong, M. M., Gujaria-Verma, N., Ramsay, L., Yuan, H. Y., Caron, C., Diapari, M., Vandenberg, A., & Bett, K. E., (2015). Classification and characterization of species within the genus lens using genotyping-by-sequencing (GBS). PlosONE 10 (3), E0122025. https://doi.org/10.1371/journal.pone.0122025
• Zhao, D., Sapkota, M., Lin, M., Beil, C., Sheehan, M., Greene, S., & Irish, B. M. (2024). Genetic diversity, population structure, and taxonomic confirmation in annual medic (Medicago spp.) collections from Crimea, Ukraine. Frontiers in Plant Science, 15, 1339298. https://doi.org/10.3389/fpls.2024.1339298