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. 2024 Oct 12;164(5-6):276–283. doi: 10.1159/000541706

Fluorescence in situ Hybridization Analysis of Oligonucleotide 5S Ribosomal DNA, 45S Ribosomal DNA, and (TTTAGGG)3 Locations in Gloriosa superba L.

Hongyou Zhao a,b, Duo Wang c,✉, Haitao Li a,b,✉, Shuang Li a,b,✉, Yanfang Wang a,b,✉, Anshun Xu a,b, Chunyong Yang a,b, Ge Li a,b,✉, Yanqian Wang a,b,✉, Lixia Zhang a,b,✉
PMCID: PMC11825083  PMID: 39396512

Abstract

Introduction

Gloriosa superba L. is a horticulturally and medicinally important plant native to Africa. However, the few cytogenetic studies of the species are mainly focused on chromosome counting and chromosome morphology-based karyotyping. Fluorescence in situ hybridization (FISH) is a powerful tool for the detection of DNA repetitive elements in a specific region of a chromosome.

Methods

Here, detailed karyotypes of G. superba were constructed by FISH using 5S and 45S rDNAs, and telomeric repeat (TTTAGGG)3 oligonucleotides.

Results and Conclusion

Twenty-two chromosomes were observed. Two 5S rDNA hybridization signals were detected in the proximal regions of the short arms of one pair of chromosomes, which were adjacent to the (TTTAGGG)3 terminal signals. Four 45S rDNA signals were detected near the centromere region of the short arm of the four chromosomes, but one of these was very weak and almost undetectable compared to the others. Telomeric repeat hybridization signals were distributed at the terminal region of each chromosome. The chromosomes displayed were intact, and the chromosome counts were accurate. Chromosome length ranged from 3.46 to 9.31 μm. These results will facilitate the cytogenetic mapping of other major repeats, thus contributing to an improved understanding of the G. superba genome structure and evolutionary history.

Keywords: Gloriosa, Fluorescence in situ hybridization, Oligonucleotide probes, Ribosomal DNA, Telomere

Introduction

Gloriosa superba L. (flame lily, glory lily, or climber lily), belonging to the Colchicaceae family, is a perennial tuberous herbaceous climbing plant with brilliant wavy-edged yellow and red flowers that appear from July to August every year. It is a native of tropical Africa and grows naturally in many parts of tropical Asia such as India, Burma, Sri Lanka, Malaysia, and Bangladesh [1, 2]. In China, it is only found in southern Yunnan [3]. The flame lily is the national flower of Zimbabwe and the state flower of Tamil Nadu in India [4]. As a consequence of its floral beauty, it is used as an ornamental plant by plant lovers worldwide [5].

Despite its showy appearance (flowers), all parts of the plant, especially the tubers and seeds, are extremely toxic and cause fatalities if consumed due to the presence of two colchicine and gloriosine alkaloids [6–8]. However, the flame lily is also an important medicinal plant due to the presence of these alkaloids. This plant is widely used to cure various human ailments in several indigenous systems of medicine. It has analgesic, anti-inflammatory [9], antimicrobial and anticancer [10], mutagenic [11], anthelmintic [12], antioxidant [13], and antithrombotic [14] potential. In addition, it can be used in the treatment of intestinal worms, gout, arthritis [5], snake bites, skin problems [15], etc.

Unfortunately, the natural resources of the flame lily have been overexploited from the wild to satisfy the demand for colchicine in national and international drug markets. G. superba is on the verge of extinction, is one of the endangered species among the most valued medicinal plants, and has an entry in the Red Data Book by the International Union for Conservation of Nature [1, 4, 16]. Therefore, it is urgent to protect the germplasm resources of G. superba. Authentic identification of the genetic composition of the species is very important before conservation [2]. Karyotype analysis is a stable and reliable method that can not only reveal cytological characteristics but also provide basic information about genetic composition. Many earlier studies have been conducted on the karyotype of G. superba, but most are limited to chromosome count [17–20] and do not provide sufficient details of the genetic makeup.

Fluorescence in situ hybridization (FISH) is a powerful tool for plant karyotype analysis, which helps provide information on the distribution of repetitive elements on chromosomes, thereby revealing patterns of genetic composition and karyotype variation [21–24]. Ribosomal DNA (rDNA) and telomeric repeats, which are two common tandem repeats, are preferred as chromosomal markers for the analysis and construction of chromosome karyotype maps in a number of plant species, such as Helianthus species [25], Triticum monococcum [26], Araliaceae species [27], Fraxinus mandshurica and F. quadrangulate [28], Hibiscus hamabo [29], Hippophae rhamnoides [30], Dracaena species [31], and Fabaceae species [32]. Thus, it seems that FISH using rDNA and telomeric repeat as probes can provide quite satisfactory, detailed, and critical chromosome analysis. For instance, FISH can be used to determine the amount and sites of rDNA and telomeric repeats in chromosomes or disclose the structural integrity of chromosome ends. In the present study, molecular cytogenetic information related to chromosome number, the distribution of rDNA, and telomeric repeats in the genome of G. superba were determined in karyotype analysis using three oligonucleotide probes: 5S rDNA, 45S rDNA, and telomeric repeats.

Materials and Methods

Plant Material

Ten seedlings of G. superba were collected from the Yunnan Branch of Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences & Peking Union Medical College, and then germinated under a 12-h photoperiod (light/dark temperature 28/24°C). When the roots reached 1–2 cm, the root tips were treated with 1.0 MPa nitrous oxide gas for 2 h, fixed in glacial acetic acid for 10 min, transferred into 75% (v/v) ethanol, and then stored at −20°C before preparing slides [33].

Chromosome and Probe Preparation

Treated root tips were washed twice with ddH2O. The meristematic zone (∼1 mm) of the root tip was excised and digested with cellulase and pectinase (4:2) for 50 min at 37°C. Next, the enzyme mixture on the meristems was cleaned by ddH2O and then washed with ethyl alcohol. All ethyl alcohol was removed, and 20 μL glacial acetic acid was added to each meristem to prepare a suspension. The mixture (10 μL) was dropped on one new slide as described by Komuro et al. [34]. The slides were examined using an Olympus CX23 Microscope (Olympus, Japan). Slides with well-spread metaphase chromosomes were stored at −20°C until use.

Three oligonucleotides were used in this work; they included TAMRA (6-carboxytetramethylrhodamine)-labeled oligonucleotides 5S rDNA (5′-CTG​ATG​GGA​TCC​GGT​GCT​TT-3′), FAM (6-carboxyfluorescein)-labeled oligonucleotides 45S rDNA (5′-TCG​TAA​CAA​GGT​TTC​CGT​AG-3′) [31], and TAMRA (6-carboxytetramethylrhodamine)-labeled Arabidopsis-type telomeric repeat (TTTAGGG)3 [35]. All oligonucleotides were synthesized by Tsingke Biotechnology Co., Ltd. (Beijing, China).

FISH and Karyotype Analysis

FISH using multiple probes was performed as described by Huang et al. [36]. The slides with a good splitting phase were denatured in 70% alcohol + NaOH (0.15 mol/L) for 6 min at 42°C and dehydrated in a 70, 95, and 100% ethyl alcohol series, each for 5 min, followed by air-drying. The hybridization buffer (10 μL, including 0.3 μL of each probe, 4.85 μL of 2×SSC, and 4.85 μL of 1×TE) was denatured at 85°C for 5 min and then transferred to −20°C 100% alcohol for 10 min, sequentially dropped on to chromosomes, and incubated for 2 h at 37°C. After hybridization, the slides were washed with 2×SSC twice and then washed with ddH2O twice at room temperature and air-dried. Chromosomes were counterstained with 4,6-diamidino-2-phenyl-indole (DAPI; Vector Laboratories, Inc., Burlingame, CA, USA). The hybridization signals were visualized with an Olympus BX-53 Microscope (Olympus, Tokyo, Japan). The images were captured with a Photometric SenSys Olympus DP73 CCD camera (Olympus), and image analysis was conducted with Photoshop V7.0 (Adobe Systems Inc., San Jose, CA, USA). After capturing images, the slides were recycled and then stored at −20°C for the next FISH [34].

At least 20 cells with well-separated metaphase chromosomes were observed to determine the chromosome number. At least three best spreads were used for the analysis of karyotype and signal patterns. The length of each chromosome was calculated using Photoshop V7.0 (Adobe Systems Inc., San Jose, CA, USA). Finally, all chromosomes were aligned by length, from the longest to the shortest.

Results

Chromosome Karyotype

High-quality somatic chromosome spreads of G. superba from root meristems were successfully prepared by using an enzyme digestion technique [34]. The mitotic metaphase chromosomes of the species detected using 5S rDNA, 45S rDNA, and (TTTAGGG)3 are illustrated in Figure 1. DAPI-stained images showed that the metaphase chromosome number of G. superba was 2n = 22 (Fig. 1, 2). To display the karyotype details clearly, each chromosome of G. superba was cut from the images in Figure 1 and aligned with the image in Figure 2 based on its centromere, length, probe signal pattern, and chromosome segregation. The chromosome length ranges in the three G. superba spreads were 3.63–8.78 μm (Fig. 2a), 3.57–9.31 μm (Fig. 2b), and 3.46–8.70 μm (Fig. 2c), respectively. The total lengths of the chromosomes were 111.58 (Fig. 2a), 111.66 (Fig. 2b), and 109.00 μm (Fig. 2c). G. superba was found to possess metacentric and sub-metacentric chromosomes, with the former greatly outnumbering the latter (Fig. 2). The first and second chromosomes were two large metacentric chromosomes, comparatively longer than the other 20 chromosomes (Fig. 2). The karyotype type of the species was 2B. The karyotype asymmetry index in each of the three spreads was 56.52%, 56.86%, and 56.66%, and the ratio of the longest to shortest chromosomes was 2.42, 2.60, and 2.51, respectively.

Fig. 1.

Fig. 1.

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a–c Somatic metaphase chromosomes of G. superba after FISH using 5S rDNA (red), 45S rDNA (green), and Arabidopsis-type telomere ((TTTAGGG)3) (red) oligonucleotide probes. a, b and c DAPI stained metaphase chromosome spreads (blue) (white arrows indicate the NOR sites); a1, b1, c1, a2, b2 and c2 45S rDNA signals (green), the chromosomes with a weak 45s rDNA signal are marked as green arrows; a2, b2 and c2 5S rDNA signals (red); a3, b3 and c3 (TTTAGGG)3 signals (red). All scale bars, 10 μm. NOR, nucleolus organizer region.

Fig. 2.

Fig. 2.

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FISH karyotyping of G. superba through hybridization with 5S rDNA (red signals, a, b and c), 45S rDNA (green signals, a, b and c), and Arabidopsis-type telomere ((TTTAGGG)3) (red signals, a′, b′ and c′) oligonucleotides. All chromosomes are captured from Fig. 1. For example, the chromosomes in a and a′ are from Figure 1a1, a2, and a3, respectively. Because weak 45S rDNA signals were easily obscured after image composition, chromosomes (orange) with the hybridization signal are included as show in a, b and c. The chromosomes were aligned by length (from the longest to shortest) and signal pattern (a–c′). The left/right and bottom numbers indicate chromosome lengths and chromosome counts (a, b and c), respectively.

rDNA and Arabidopsis-Type Telomere Repeat Distribution Patterns

To determine the number and chromosomal distributions of rDNA on the G. superba chromosome spreads, 5S and 45S rDNA oligonucleotide probes were applied using FISH. The number and distribution of the 5S and 45S rDNA hybridization signals were different in this species. Two 5S and four 45S rDNA hybridization signals were observed on six chromosomes: one site of a 5S locus and two sites of 45S loci were located on three different pairs of homologous chromosomes (Fig. 1). Weak 5S rDNA hybridization signals were located at the proximal regions of the short arms on two chromosomes (11/12) (Fig. 1, 2), which were adjacent to the (TTTAGGG)3 terminal hybridization signals on these two chromosomes, which were a pair of metacentric chromosomes (Fig. 2a–c′). Two types of hybridization signal patterns were found using 45S rDNA based on FISH signal intensities (Fig. 1, 2). Type I 45S rDNA clearly showed a strong FISH signal, which was located interstitially near the centromere region on the short arm of the three chromosomes (13/14/15). Type II showed a weaker FISH signal than the former. It was located toward the centromere region on the short arm of the chromosome (16). Furthermore, the larger nucleolus organizer region of the former was clearly distinguishable and could easily be identified after DAPI staining, whereas the latter was barely observable (Fig. 1a–c). DAPI images revealed that three chromosomes (13/14/15) were sub-metacentric chromosomes and one chromosome (16) was a metacentric chromosome (Fig. 2), where the centromere positions are clearly distinguished. The three sub-metacentric chromosomes (13/14/15) were significantly longer than the metacentric chromosome (16) in length.

Additionally, FISH analysis of G. superba was performed using telomere repeats (TTTAGGG)3 as probe. The probe produced clear hybridization signals at the end of each chromosome (Fig. 1, 2). This result indicated that the chromosomes were intact and the chromosome number was accurate.

Discussion

Karyotype Analysis

Chromosome number, size, and morphological characteristics are important components of karyotypes. In our study, 22 chromosomes were observed in G. superba. Our results for G. superba (2n = 22) confirm the chromosome counts reported earlier [2, 18–20, 37–40]. Other chromosome counts for this species have also been reported such as 2n = 44 [41], 2n= 56 [42], 2n = 77 [43], 2n = 88 [44], and 2n = 90 [45]. Our results for G. superba (2n = 22) did not support these findings. The genus Gloriosa with a genetic base x = 11 includes diploids such as G. superba, G. lutea, and G. plantii (2n = 2x = 22), and polyploids, G. carsonii, G. virescens, and G. richmondensis (2n = 4x = 44) and G. rothschildiana, G. latifolia, and G. magnifica (2n = 8x = 88) [5]. Thus, the chromosome numbers 2n = 44, 77, and 88 for G. superba can be interpreted as tetraploid, septaploid, and octaploid derived from x = 11, whereas 2n= 56 and 90 might be a case of aneuploidy. G. superba exhibits intraspecific polyploidy and aneuploidy at polyploid levels. However, these different chromosome counts may also be the result of botanical misidentification or outdated chromosome preparation techniques [46].

The metaphase chromosome length range of G. superba was 3.46–9.31 μm in the current study. In previous works, the chromosome lengths of this species were reported to be 2.84–9.67 μm [18] and 3.28–7.83 μm [2], which are similar to the lengths we obtained. The total lengths of the chromosomes were 109.00–111.66 μm, which is within the range of 98.64–129.06 μm previously reported by Vijayavalli and Mathew [18]. There were two types of chromosomes in G. superba: metacentric and sub-metacentric. The first and second chromosomes were two large metacentric chromosomes. Here, we did not observe one large metacentric chromosome and another large sub-metacentric chromosome between them as described by Sultana et al. [2]. The karyotype asymmetry index was 56.52% to 56.86%, and the ratio of the longest to shortest chromosomes was 2.42 to 2.60, which indicated karyotype asymmetry in this species.

Roles of rDNA and Arabidopsis-Type Telomere Repeats

FISH can refine cytogenetic karyotyping and can identify different chromosome markers for physical mapping [47, 48]. A combination of 5S and 45S rDNA is widely used as chromosome markers for routine FISH analysis in many plant species, e.g., Allium cepa L. [49], Rosa [47], Senna [48], and Dracaena [31]. Using FISH, we found that the 5S rDNA was quite conserved. It was located only at the proximal regions of two chromosomes. The physical positions of 45S rDNA loci were also relatively conservative. Hybridization signals involving 45S rDNA were located interstitially near the centromere region on four chromosomes. However, compared with 5S, 45S rDNA showed polymorphism in signal intensity. One 45S rDNA signal was very weak or almost invisible compared to the other three; i.e., 45S rDNA locus had an unbalanced copy number in one homolog. A similar situation of apparent 45S rDNA locus loss was also observed in Paphiopedilum [50], Allium cepa L. [49], Brassica rapa L., B. napus L. [51], and Senna [48]. Tandem repeats undergo frequent recombination and unequal crossing over that lead to unbalanced copy numbers of the whole tandem repeat [52, 53]. Thus, the change in locus size could be explained by the recombination of repetitive DNAs. Importantly, we could not ignore the fact that the much shorter 45S rDNA arrays were not detected due to the limited resolution of the FISH technique.

(TTTAGGG)n, an Arabidopsis-type telomere repeat sequence, is a typical chromosome maker that is useful for characterizing basic telomeric repeats in higher plant phyla [54]. Arabidopsis-type telomeric repeats are present in nearly all plants except the 16 species in 12 families of Asparagales and other species in the genera Allium (Liliaceae), Nothoscordum, and Tulbaghia (Alliaceae) previously studied [55]. The typical telomere sequence is often localized at chromosomal termini in many plant species, such as Aralia elata (Miq.) Seem., Dendropanax morbiferus H. Lév., Eleutherococcus sessiliflorus (Rupr. Et Maxim.) Seem., Kalopanax septemlobus (Thunb. ex A.Murr.) Koidz. [27], Citrus sinensis × Poncirus trifoliata [56], and Dracaena Vand. ex L. species [31]. However, the Arabidopsis-type telomere sequence was occasionally detected at para- or peri-centromeric and interstitial regions of chromosomes in Arabidopsis thaliana (L.) Heynh. [35], Rosa L. [47], Senna [48], and tomato [57]. These unconventional non-telomeric blocks or interstitial telomere repeats are possibly relics of chromosome rearrangement during evolution. Here, our results showed that no interstitial telomere signals were observed on the chromosomes of G. superba.

Conclusions

To our knowledge, this is the first oligo-FISH karyotype report in G. superba as well as within the genus Gloriosa. Our results provide accurate chromosome information and efficient chromosome identification to better understand the genome structure of this species. As several chromosomes remain to be identified, further studies are needed to develop repeat-based cytogenetic markers to distinguish individual chromosomes. It would be useful to further refine the karyotypes of the glory lily and to trace the history of its genome structure and evolution.

Acknowledgments

The authors thank Zhao Laibin and Liu Juncheng, for experiment method instruction.

Statement of Ethics

An ethics statement was not required for this study type, and no human or animal subjects were used.

Conflict of Interest Statement

The authors declare no competing financial interests.

Funding Sources

This research was supported by the CAMS Innovation Fund for Medical Sciences (CIFMS) (No. 2021-I2M-1-032), the Yunnan Provincial Science and Technology Major Project of Biomedicine (No. 202105AG070011--YZS-24), the Yunnan Provincial Science and Technology Talents and Platform Project (No. 202205AF150071, No. 202105AD160054), and the Selection Special Programme of Yunnan Province High-level Technological Talents and Innovative Teams (No. 202405AS350020).

Author Contributions

H.Z. and D.W. contributed equally to this work. H.Z., D.W., Y.W. (Yanqian Wang), and L.Z designed the study, analyzed the data, and wrote the manuscript. H.Z., D.W., H.L., S.L., and Y.W (Yanfang Wang) performed the experiments. C.Y., G.L., and A.X. discussed the manuscript.

Funding Statement

This research was supported by the CAMS Innovation Fund for Medical Sciences (CIFMS) (No. 2021-I2M-1-032), the Yunnan Provincial Science and Technology Major Project of Biomedicine (No. 202105AG070011--YZS-24), the Yunnan Provincial Science and Technology Talents and Platform Project (No. 202205AF150071, No. 202105AD160054), and the Selection Special Programme of Yunnan Province High-level Technological Talents and Innovative Teams (No. 202405AS350020).

Data Availability Statement

The authors confirm that the data supporting the findings of this study are available within the article and its supplementary materials.

References

  • 1. Jana S, Shekhawat GS. Critical review on medicinally potent plant species: Gloriosa superba. Fitoterapia. 2011;82(3):293–301. [DOI] [PubMed] [Google Scholar]
  • 2. Sultana SS, Dash CK, Alam SS, Hassan MA. Karyotype analysis and report on B-chromosome in Gloriosa superba L. by differential staining. Nucleus. 2018;62(1):31–8. [Google Scholar]
  • 3. Wu ZY, Raven PH, Hong DY, editors. Flora of China. 2nd edn.Beijing & St. Louis: Missouri Botanical Garden Press, Science Press; 2000. [Google Scholar]
  • 4. Mahajan R. Gloriosa superba L. : An endangered medicinal plant. Hort Flora Res Spectr. 2015;4(2):168–71. [Google Scholar]
  • 5. Padmapriya S, Rajamani K, Sathiyamurthy VA. Glory lily (Gloriosa superba L.)-A review. Int J Curr Pharm Rev Res. 2016;7(1):43–9. [Google Scholar]
  • 6. Muthukrishnan SD, Annapoorani Subramaniyan AS. Phytochemical constituents of gloriosa superba seed, tuber and leaves. Res J Pharm Biol Chem Sci. 2012;3(3):111–7. [Google Scholar]
  • 7. Phatak RS, Hegde LN. Glory lily (Glorosa superba L.): an important medicinal plant crop: a review. Hort Flora Res Spectr. 2014;3(3):282–7. [Google Scholar]
  • 8. Ashokkumar K. Gloriosa superba (L.): a brief review of its phytochemical properties and pharmacology. Int J Pharm Phytochem Res. 2015;7(6):1190–3. [Google Scholar]
  • 9. Deshmukh P, John J, Fernandes J, Nandgude T, Niphade S, Savla A. Analgesic and anti-inflammatory activities of the hydroalcoholic extract from Gloriosa superba Linn. Int J Green Pharm. 2009;3(3):215–9. [Google Scholar]
  • 10. Budhiraja A, Nepali K, Kaul S, Dhar KL. Antimicrobial and cytotoxic activities of fungal isolates of medicinal plant Gloriosa superba. Int J Recent Adv Pharm Res. 2012;2(1):37–45. [Google Scholar]
  • 11. Hemaiswarya S, Raja R, Anbazhagan C, Thiagarajan V. Antimicrobial and mutagenic properties of the root tubers of Gloriosa superba Linn. (Kalihari) Pak J Bot. 2009;41(1):293–9. [Google Scholar]
  • 12. Suryavanshi S, Rai G, Malviya N. Evaluation of anti-microbial and anthelmintic activity of Gloriosa Superba tubers. Adv Res Pharm Biol. 2012;2(1):45–52. [Google Scholar]
  • 13. Jagtap S, Satpute R. Phytochemical screening, antioxidant, antimicrobial and flavonoid analysis of Gloriosa superba linn. Rhizome extracts. J Acad Ind Res. 2014;3(6):247–54. [Google Scholar]
  • 14. Kee NLA, Mnonopi N, Davids H, Naudé RJ, Frost CL. Antithrombotic/anticoagulant and anticancer activities of selected medicinal plants from South Africa. Afr J Biotechnol. 2008;7(3):217–23. [Google Scholar]
  • 15. Kavina JGR, Panneerselvam R. Gloriosa superba linn: a medicinally important plant. Drug Invent Today. 2011(6):69–71. [Google Scholar]
  • 16. Lal HS, Mishra PK. Gloriosa Superba: an Endangered plant spotted for the first time from forest of Tpchanchi, Hazaribag (Jharkhand) India. Sci Res Rep. 2011;1(2):61–4. [Google Scholar]
  • 17. Lemattre P. Chromosomic number of some gloriosa. Acta Hortic. 1980;109:357–60. [Google Scholar]
  • 18. Vijayavalli B, Mathew PM. Karyomorphology of four morphotypes of Gloriosa superba L. From South India. Cytologia. 1990;55(4):531–3. [Google Scholar]
  • 19. Amano J, Kuwayama S, Mizuta Y, Nakano M, Godo T, Okuno H. Morphological characterization of three intergeneric hybrids among Gloriosa superba ‘lutea’, Littonia modesta, and Sandersonia aurantiaca (Colchicaceae). HortScience. 2008;43(1):115–8. [Google Scholar]
  • 20. Ramesh A. Chromosomal study and medicinal uses in Gloriosa superba L. Int Res J Biol Sci. 2015;4(5):4–6. [Google Scholar]
  • 21. Fu SL, Chen L, Wang YY, Li M, Yang ZJ, Qiu L, et al. Oligonucleotide probes for ND-FISH analysis to identify rye and wheat chromosomes. Sci Rep. 2015;5(1):10552–7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22. Wang DR, Pei D, Pei ZY, Zhuang LF, Qi ZJ. Development and application of high resolution karyotypes of wheat “Chinese spring” aneuploids. Acta Agron Sin. 2017;43(11):1575–87. [Google Scholar]
  • 23. Zhao LB, Ning SZ, Yi YJ, Zhang LQ, Yuan ZW, Wang JR, et al. Fluorescence in situ hybridization karyotyping reveals the presence of two distinct genomes in the taxon Aegilops tauschii. BMC Genomics. 2018;19(1):3–9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24. Luo XM, He ZJ, Liu JC, Wu HY, Gong X. FISH mapping of telomeric and non-telomeric (AG3T3)3 reveal the chromosome numbers and chromosome rearrangements of 41 woody plants. Genes. 2022;13(7):1239–24. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25. Vanzela ALL, Ruas CF, Oliveira MF, Ruas PM. Characterization of diploid, tetraploid and hexaploid Helianthus species by chromosome banding and FISH with 45S rDNA probe. Genetica. 2002;114(2):105–11. [DOI] [PubMed] [Google Scholar]
  • 26. Megyeri M, Mikó P, Farkas A, Molnár-Láng M, Molnár I. Cytomolecular discrimination of the Am chromosomes of Triticum monococcum and the A chromosomes of Triticum aestivum using microsatellite DNA repeats. J Appl Genet. 2017;58(1):67–70. [DOI] [PubMed] [Google Scholar]
  • 27. Zhou HC, Pellerin RJ, Waminal NE, Yang TJ, Kim HH. Pre-labelled oligo probe-FISH karyotype analyses of four Araliaceae species using rDNA and telomeric repeat. Genes Genomics. 2019;41(7):839–47. [DOI] [PubMed] [Google Scholar]
  • 28. Islam-Faridi N, Mason ME, Koch JL, Nelson D. Cytogenetics of Fraxinus mandshurica and F. quadrangulata: ploidy determination and rDNA analysis. Tree Genet Genomes. 2020;16(1):26–7. [Google Scholar]
  • 29. Sakhanokho HF, Islam-Faridi N, Babiker EM, Nelson CD, Stringer SJ, Adamczyk JJ Jr. Determination of nuclear DNA content, ploidy, and FISH location of ribosomal DNA in Hibiscus hamabo. Sci Hortic. 2020;264:109167–7. [Google Scholar]
  • 30. Luo XM, Liu JC, He ZJ. Oligo-FISH can identify chromosomes and distinguish hippophaë rhamnoides L. Taxa. Genes. 2022;13(2):195–16. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31. Zhao HY, Li S, Yang CY, Li G, Wang YF, Peng JM, et al. FISH-based karyotype analyses of four Dracaena species. Cytogenet Genome Res. 2021;161(5):272–7. [DOI] [PubMed] [Google Scholar]
  • 32. He ZJ, Zhang W, Luo XM, Huan J. Five Fabaceae karyotype and phylogenetic relationship analysis based on oligo-FISH for 5S rDNA and (AG3T3)3. Genes. 2022;13(5):768–10. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33. Kato A. Air drying method using nitrous oxide for chromosome counting in maize. Biotech Histochem. 1999;74(3):160–6. [DOI] [PubMed] [Google Scholar]
  • 34. Komuro S, Endo R, Shikata K, Kato A. Genomic and chromosomal distribution patterns of various repeated DNA sequences in wheat revealed by a fluorescence in situ hybridization procedure. Genome. 2013;56(3):131–7. [DOI] [PubMed] [Google Scholar]
  • 35. Richards EJ, Goodman HM, Ausubel FM. The centromere region of Arabidopsis thaliana chromosome 1 contains telomere-similar sequence. Nucleic Acids Res. 1991;19(12):3351–7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36. Huang XY, Zhu MQ, Zhuang LF, Zhang SY, Wang JJ, Chen XJ, et al. Structural chromosome rearrangements and polymorphisms identified in Chinese wheat cultivars by high-resolution multiplex oligonucleotide FISH. Theor Appl Genet. 2018;131(9):1967–86. [DOI] [PubMed] [Google Scholar]
  • 37. Bolkhovskikh ZV, Grip VG, Zakhar’Eva OI, Matveeva TS, AI E, Fedorov AA. Chromosome numbers of flowering plants. Bot Inst.; 1969. [Google Scholar]
  • 38. Mehra PN, Sachdeva SK. Cytological observations on some W. Himalayan monocots II. Smilacaceae, liliaceae and trilliaceae. Cytologia. 1976;41(1):5–22. [Google Scholar]
  • 39. Goldblatt P. Index to plant chromosome numbers 1979–1981. 1984. [Google Scholar]
  • 40. Vijayavalli B, Mathew PM. Cytology of species of gloriosa linn. J Indian Bot Soc. 1992;71:29–32. [Google Scholar]
  • 41. Narain P. Chromosomal mosaicism in the microsporocytes of Gloriosa. Cytologia. 1980;45(1/2):271–9. [Google Scholar]
  • 42. Nwofia GE, Madu OA, Eneobong EE. Studies on the chromosomes of tropical lilies 2: karyotype of Gloriosa superba lindle (liliaceae). J Appl Chem Agric Res. 2008;7(1). [Google Scholar]
  • 43. Kishimoto T, Nakazawa D, Sato T, Saito T, Amano J, Kuwayama S, et al. Horticultural characterization of an intergeneric hybrid between a septaploid gloriosa cultivar and Sandersonia aurantiaca. Acta Hortic. 2014;1025:51–6. [Google Scholar]
  • 44. La-Cour LF. Heterochromatin and the organization of nucleoli in plants. Heredity. 1951;5(1):37–50. [DOI] [PubMed] [Google Scholar]
  • 45. Sharma AK, Sharma A. An investigation of the cytology of some species of Liliaceae. Genet Iber. 1961;13:25–8. [Google Scholar]
  • 46. Guerra M. Chromosome numbers in plant cytotaxonomy: concepts and implications. Cytogenet Genome Res. 2008;120(3–4):339–50. [DOI] [PubMed] [Google Scholar]
  • 47. Kirov IV, Van Laere K, Van Roy N, Khrustaleva LI. Towards a FISH-based karyotype of Rosa L. (Rosaceae). Comp Cytogenet. 2016;10(4):543–54. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 48. Pellerin RJ, Waminal NE, Kim HH. FISH mapping of rDNA and telomeric repeats in 10 Senna species. Hortic Environ Biotechnol. 2019;60(2):253–60. [Google Scholar]
  • 49. Mancia FH, Sohn SH, Ahn YK, Kim DS, Kim JS, Kwon YS, et al. Distribution of various types of repetitive DNAs in Allium cepa L. based on dual color FISH. Hortic Environ Biotechnol. 2015;56(6):793–9. [Google Scholar]
  • 50. Lan T, Albert VA. Dynamic distribution patterns of ribosomal DNA and chromosomal evolution in Paphiopedilum, a lady’s slipper orchid. BMC Plant Biol. 2011;11:126–15. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 51. Waminal NE, Perumal S, Lee J, Kim HH, Yang TJ. Repeat evolution in Brassica rapa (AA), B. oleracea (CC), and B. napus (AACC) genomes. Plant Breed. 2016;4(2):107–22. [Google Scholar]
  • 52. Dover GA. Linkage disequilibrium and molecular drive in the rDNA gene family. Genetics. 1989;122(1):249–52. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 53. Liao DQ. Concerted evolution: molecular mechanism and biological implications. Am J Hum Genet. 1999;64(1):24–30. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 54. Fuchs J, Brandes A, Schubert I. Telomere sequence localization and karyotype evolution in higher plants. Pl Syst Evol. 1995;196(3–4):227–41. [Google Scholar]
  • 55. Adams SP, Hartman TPV, Lim KY, Chase MW, Bennett MD, Leitch IJ, et al. Loss and recovery of Arabidopsis-type telomere repeat sequences 5′-(TTTAGGG)n-3′ in the evolution of a major radiation of flowering plants. Proc Biol Sci. 2001;268(1476):1541–6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 56. Deng H, Tang G, Xu N, Gao Z, Lin L, Liang D, et al. Integrate and d karyotypes of diploid and tetraploid carrizo citrange (Citrus sinensis L. Osbeck x Poncirus trifoliata L. Raf.) as determined by sequential multicolor fluorescence in situ hybridization with tandemly repeated DNA sequences. Front Plant Sci. 2020;11:569–78. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 57. Presting GG, Frary KP, Tanksley SD. Telomere-homologous sequence occur near the centromeres of many tomato chromosomes. Mol Gen Genet. 1996;251:526–31. [DOI] [PubMed] [Google Scholar]

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Data Availability Statement

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