First Trials of Genome Analyses in Some Onobrychis Species using Dot-Blot and Genomic in situ Hybridization Techniques

Abstract

The origin and genome composition of tetraploid cultivated Onobrychis viciifolia (2n = 4x = 28) were analyzed using dot-blot and genomic in situ hybridization (GISH) techniques. Dot-blot hybridization was used to find a genomic affinity between O. viciifolia and 16 diploid Onobrychis species. The hypothesis on the origin of the O. viciifolia was tested using GISH. Dot-blot analyses suggested a genomic affinity between O. viciifolia and four diploid Onobrychis species (O. kachetica, O. supina, O. pallasii, and O. vaginalis). Hybridization signals were observed on O. viciifolia chromosomes when gDNA of O. kachetica, O. supina, O. pallasi, and O. hypargyrea were used as probes. However, the observed chromosomal distribution of hybridization signals did not resemble GISH results. The observed signals colocalized with 35S rDNA or dispearse signals on all chromosomes were observed depending on the probe. Further investigations using more comprehensive and comparative analysis with both coding and repetitive DNA regions may provide a better understanding of the genome composition and evolution of O. viciifolia.

Keywords

• Onobrychis
• Dot-blot hybridization
• GISH
• FISH

References

1. C. H. Carbonero, I. Mueller-Harvey, T. A. Brown, L. Smith, Sainfoin (Onobrychis viciifolia): A beneficial forage legume, Plant Genetic Resources: Characterization and Utilization 9 (1) (2011) 70–85.
2. C. H. Carbonero, F. Carbonero, L. M. J. Smith, T. A. Brown, Phylogenetic characterisation of Onobrychis species with special focus on the forage crop Onobrychis viciifolia Scop, Genetic Resources and Crop Evolution 59 (2012) 1777–1788.
3. T. Sutcu, B. B. Bilgen, M. Tuna, Analysis of genetic diversity among Onobrychis accessions with high agronomic performance by simple sequence repeat (SSR) markers, Molecular Biology Reports 49 (2022) 5659–5668.
4. B. Yıldız, B. Çıplak, E. Aktoklu, Fruit morphology of sections of the genus Onobrychis Miller (Fabaceae) and its phylogenetic implications, Israel Journal of Plant Sciences 47 (4) (1999) 269–282.
5. G. I. Sirjaev, Onobrychis Generis Revisio Critica, Pars Prima; Faculte’ des Sciences de I’Universite Masaryk: Brno, Brno, 1925.
6. E. C. Yilmaz, B. B. Bilgen, M. Tuna, Evaluation of the genetic structure of some accessions belonging to Onobrychis spp. using microsatellite DNA markers, Journal of Tekirdag Agricultural Faculty 20 (2) (2023) 399-409.
7. M. Mora-Ortiz, L. Smith, Onobrychis viciifolia; a comprehensive literature review of its history, etymology, taxonomy, genetics, agronomy and botany, Plant Genetic Resources 16 (5) (2018) 403–418.
8. A. Amirahmadi, S. Kazempour-Osaloo, A. Kaveh, A. A Maassoumi, R. Naderi, The phylogeny and new classification of the genus Onobrychis (Fabaceae-Hedysareae): Evidence from molecular data, Plant Systematics and Evolution 302 (2016). 1445–1456.

9. C. H. Carbonero, F. Carbonero, L. M. J. Smith, T. A. Brown, Cytological characterisation of the underutilized forage crop Onobrychis viciifolia Scop. and other members of the Onobrychis genus, Genetic Resources and Crop Evolution 60 (7) (2013) 1987–1996.
10. M. M. Abou-El-Enain, Chromosomal criteria and their phylogenetic implications in the genus Onobrychis Mill. sect. Lophobrychis (Leguminosae), with special reference to Egyptian species, Botanical Journal of Linnean Society 139 (4) (2002) 409–414.
11. H. Hoşgören, Total numbers of chromosome numbers in species of Onobrychis Miller (Fabaceae) in Southeastern Anatolia region, Biotechnology Biotechnoligal Equipment 20 (2) (2006) 57–61.
12. H. H. S. Mohsen, M. Z. Nasab, Cytotaxonomy of some Onobrychis (Fabaceae) species and populations in Iran, Caryologia 63 (2010) 18–31.
13. S. E. Akçelik, S. Avcı, S. Uzun, C. Sancak, Karyotype analysis of some Onobrychis (sainfoin) species in Turkey, Archieves of Biological Sciences 64 (2012) 567–571.
14. M. Ranjbar, F. Hajmoradi, R. Karamian, An overview on cytogenetics of the genus Onobrychis (Fabaceae) with special reference to O. sect. Hymenobrychis from Iran, Caryologia 65 (2012) 187–198.
15. G. Yücel, A. Betekhtin, E. Cabi, M. Tuna, R Hasterok, B. Kolano, The chromosome number and rDNA loci evolution in Onobrychis (Fabaceae), International Journal of Molecular Sciences 23 (19) (2022) 11033 17 pages.
16. R. Volkov, F. Medina, U. Zentgraf, V. Hemleben, Molecular Cell Biology: Organization and molecular evolution of rDNA, nucleolar dominance and nucleolus structure, Progress in Botany, Springer-Verlag, Berlin, 2004, Ch. 4 pp. 106–146.
17. I. Alvarez, J. F. Wendel, Ribosomal ITS sequences and plant phylogenetic inference, Molecular Phylogenetics and Evolution 29 (3) (2003) 417–434.
18. G. N. Feliner, J. A. Rossello, Better the devil you know? Guidelines for insightful utilization of nrDNA ITS in species-level evolutionary studies in plants, Molecular Phylogenetics and Evolution 44 (2) (2007) 911–919.
19. M. A. Lysak, I. Schubert, Mechanisms of chromosome rearrangements, Plant Genome Diversity Volume 2, Springer, Vienna, 2013, Ch. 9, pp. 137–147.
20. M. Te Beest, J. J. Le Roux, D. M. Richardson, A. K. Brysting, J. Suda, M. Kubesova, P. Pysek, The more the better? The role of polyploidy in facilitating plant invasions, Annals of Botany 109 (1) (2012) 19–45.
21. A. I. Bhat, G. P. Rao, Dot-Blot hybridization technique, Characterization of Plant Viruses, Springer Protocols Handbooks, New York, 2020, Ch. 34, pp. 303–321.
22. G. Robledo, G. Seijo, Characterization of the Arachis (Leguminosae) D genome using Fluorescence in situ Hybridization (FISH) chromosome markers and total genome DNA hybridization, Genetics and Molecular Biology 31(3) (2008) 717–724.
23. M. Markova, B. Vyskot, New horizons of genomic in situ hybridization, Cytogenetic and Genome Research 126 (4) (2009) 368–375.
24. B. Kolano, J. McCann, M. Orzechowska, D. Siwinska, E. Temsch, H. Weiss-Schneeweiss, Molecular and cytogenetic evidence for an allotetraploid origin of Chenopodium quinoa and C. berlandieri (Amaranthaceae), Molecular Phylogenetics and Evolution 100 (2016) 109–123.
25. C. A. Melo, G. S. Silva, M. M. Souza Establishment of the genomic in situ hybridization (GISH) technique for analysis in interspecific hybrids of Passiflora, Genetics and Molecular Research: GMR 14 (1) (2015) 2176–2188.
26. K. Emadzade, T.-S. Jang, J. Macas, A. Kovařík, P. Novák, J. Parker, H. Weiss-Schneeweiss, Differential amplification of satellite PaB6 in chromosomally hypervariable Prospero autumnale complex (Hyacinthaceae), Annal of Botany 114 (8) (2014) 1597–1608.
27. R. Hasterok, J. Draper, G. Jenkins, Laying the cytotaxonomic foundations of a new model grass, Brachypodium distachyon (L.) Beauv, Chromosome Research, 12 (2004) 397–403.
28. B. Kolano, J. McCann, M. Oskedra, M. Chrapek, M. Rojek, A. Nobis, H. Weiss-Schneeweiss, Parental origin and genome evolution of several Eurasian hexaploid species of Chenopodium (Chenopodiaceae), Phytotaxa 392 (3) (2019) 163–185.
29. I. Unfried, P. Gruendler, Nucleotide sequence of the 5.8S and 25S rRNA genes and of the internal transcribed spacers from Arabidopsis thaliana, Nucleic Acids Researches 18 (13) (1990) 4011 1 pages.
30. G. Jenkins, R. Hasterok, BAC “landing” on chromosomes of Brachypodium distachyon for comparative genome alignment, Nature Protocols 2 (2007) 88–98.
31. A. L. Tek, S. D. Kara Öztürk, High allelic diversity of the centromere-specific histone H3 (CENH3) in the legume sainfoin (Onobrychis viciifolia), Molecular Biology Reports 47 (2020) 8789–8795.
32. S. D. Kara Öztürk, Analysis of centromeric heterochromatin components by bioinformatics, molecular and cytogenetics methods in sainfoin (Onobrychis viciifolia scop.), Master Thesis Niğde Ömer Halisdemir University (2022) Niğde.
33. M. D. Sacristan, Estudios citotaxonómicos sobre el género Onobrychis (L). Adanson conreferencia especial a la citogenética de la esparceta (O. viciifolia Scop.), Zaragoza, Spain, 1965.
34. M. C. De Vicente, P. Arus, Tetrasomic inheritance of isozymes in sainfoin (Onobrychis viciaefolia Scop.), Journal of Heredity 87 (1996) 54–62.
35. M. Zarrabian, M. M. Majidi, M. H. Ehtemam, Genetic diversity in a worldwide collection of sainfoin using morphological, anatomical, and molecular markers, Crop Science 53 (6) (2013) 2483–2496.
36. K. Kempf, Self-fertilization and marker-trait associations in Sainfoin (Onobrychis viciifolia), Doctoral Dissertation ETH (2016) Zurich.
37. S. Shen, X. Chai, Q. Zhou, D. Luo, Y. Wang, Z. Liu, Development of polymorphic EST-SSR markers and characterization of the autotetraploid genome of sainfoin (Onobrychis viciifolia), PeerJ, 7:e6542 19 pages.
38. J. He, D. Tian, X. Li, X. Wang, T. Wang, Z. Wang, H. Zang, X. He, T. Zhang, Q. Yun, R. Zhang, R. Jiang., S. Jia, Y. Zhang, A chromosome-level genome assembly for Onobrychis viciifolia reveals gene copy number gain underlying enhanced proanthocyanidin biosynthesis, Communicaitions Biology 7 19 (2024).
39. A. Marasek, R. Hasterok, K. Wiejacha, T. Orlikowska, Determination by GISH and FISH of hybrid status in Lilium, Hereditas 140 (1) (2004) 1–7.
40. G. Seijo, G.I. Lavia, A. Fernandez, A. Krapovickas, D.A. Ducasse, D. J. Bertioli, E. A. Moscone, Genomic relationships between the cultivated peanut (Arachis hypogaea, Leguminosae) and its close relatives revealed by double GISH, American Journal of Botany 94 (2007) 1963–1971.
41. Z. Fredotovic, I. Samanic, H. Weiss-Schneeweiss, J. Kamenjarin, T. S. Jang, J. Puizina, Triparental origin of triploid onion, Allium x cornutum (Clementi exVisiani, 1842), as evidenced by molecular, phylogenetic and cytogenetic analyses, BMC Plant Biology 14 (2014) Article Number 24 14 pages.
42. J. F. Zoller, Y. Yang, R. G. Herrmann, U. Hohmann, Comparative genomic in situ hybridization (cGISH) analysis on plant chromosomes revealed by labelled Arabidopsis DNA, Chromosome Research 9 (2001) 357–375.
43. H. B. Ali, M. A. Lysak, I. Schubert, Genomic in situ hybridization in plants with small genomes is feasible and elucidates the chromosomal parentage in interspecific Arabidopsis hybrids, Genome 47 (5) (2004) 954–960.
44. K. Wolfe, Yesterday's polyploids and the mystery of diploidization, Nature Reviews Genetics 2 (2001) 333–341.
45. I. J. Leitch, L. Hanson, K. Y. Lim, A. Kovarik, M. W. Chase, J. J. Clarkson, A. R. Leitch, The ups and downs of genome size evolution in polyploid species of Nicotiana (Solanaceae), Annals of Botany 101 (6) (2008) 805–814.
46. T. S. Jang, H. Weiss-Schneeweiss, Formamide-free genomic in situ hybridization allows unambiguous discrimination of highly similar parental genomes in diploid hybrids and allopolyploids, Cytogenetic and Genome Research 146 (4) (2015) 325–331.