Turkish Journal of Botany Turk J Bot (2019) 43: 232-242 http://journals.tubitak.gov.tr/botany/ © TÜBİTAK Research Article doi:10.3906/bot-1807-1

Karyotype analyses of 25 Turkish taxa of Astragalus from the sections Macrophyllium, Hymenostegis, Hymenocoleus, and Anthylloidei (Fabaceae)

1 2 3 4, Esra MARTİN , Gizem İÇYER DOĞAN , Seher KARAMAN ERKUL , Halil Erhan EROĞLU * 1 Department of Biotechnology, Faculty of Science, Necmettin Erbakan University, Konya, 2 Department of Biology, Graduate School of Natural and Applied Sciences, Necmettin Erbakan University, Konya, Turkey 3 Department of Biology, Faculty of Science and Arts, , Aksaray, Turkey 4 Department of Biology, Faculty of Science and Arts, , Yozgat, Turkey

Received: 02.07.2018 Accepted/Published Online: 07.12.2018 Final Version: 07.03.2019

Abstract: The karyotype analyses and chromosomal data of 25 taxa belonging to the sections Macrophyllium, Hymenostegis, Hymenocoleus, and Anthylloidei of the genus Astragalus naturally distributed in Turkey are described. The cytogenetic data obtained were evaluated systematically. The following somatic chromosome numbers were found: 2n = 16 for A. oleaefolius, A. dipodurus, A. cephalotes, and A. yukselii; 2n = 32 for A. longifolius; and 2n = 48 for A. isauricus in the sect. Macrophyllium; 2n = 16 for A. zohrabi, A. sosnowskyi, A. velenovskyi, A. trifoliastrum, and A. uraniolimneus; 2n = 18 for A. hymenocystis; 2n = 48 for A. lagopoides, A. gueruenensis, and A. ciloensis; and 2n = 64 for A. hirticalyx in the sect. Hymenostegis; 2n = 48 for A. vaginans in the sect. Hymenocoleus; and 2n = 16 for A. szowitsii, A. ermineus, A. zederbaueri, A. anthylloides, A. halicacabus, A. chardinii, and A. wagneri; and 2n = 48 for A. surugensis in the sect. Anthylloidei. In all taxa, the karyotype results were obtained for the first time.A. szowitsii is the most symmetrical karyotype in both index values. While A. zohrabi is the most asymmetrical karyotype in intrachromosomal asymmetry, A. wagneri is the most asymmetrical karyotype in interchromosomal asymmetry.

Key words: Astragalus, asymmetry index, Fabaceae, karyotype

1. Introduction numbers were 2n = 14, 16, 22, 24, 32, 48, and 64 The genus Astragalus L. (Fabaceae) is represented by (Spellenberg, 1981; Hung, 1984; Maassoumi, 1987; Ward approximately 3000 taxa all over the world. The genus and Spellenberg, 1988; Magulaev, 1989; Liston, 1990; is spread more in arid/semiarid and cold regions of the Daviña and Gómez-Sosa, 1993; Kandemir et al., 1996; Old World with approximately 2400 taxa, North America Aytaç, 1997; Martin et al., 2008; Sheidai et al., 2009; Kazem with approximately 450 taxa, and South America with et al., 2010; Borjian et al., 2012; Rani et al., 2014). approximately 100 taxa (Keskin and Kaçar, 2013; Karaman Literature studies of taxa belonging to the genus Erkul et al., 2016; Adesso et al., 2018). Astragalus have revealed that there are too few karyotype In the flora of Turkey, the genus Astragalus is analyses of this genus. To eliminate deficiencies, represented by 64 sections and about 475 taxa with 202 somatic chromosome morphologies were identified endemic species. These species are mostly spread in the for 25 taxa belonging to the genus Astragalus that grow Irano-Turanian phytogeographical region (İlçim and naturally in Turkey, and chromosome morphology Behçet, 2016). differences were described. These taxa are A. oleaefolius DC., Many taxa of the genus Astragalus, which has an A. longifolius Lam., A. isauricus Hub.-Mur. & Matthews, important place in Turkey and the rest of the world, are A. dipodurus Bunge, A. cephalotes Banks & Sol, A. yukselii cultivated widely as medicinal plants, ornamental plants, Karaman & Aytaç, A. hymenocystis Fisch. & C.A.Mey., A. and foods, and they are used in medicine and the cosmetics lagopoides Lam., A. hirticalyx Boiss. & Kotschy, A. zohrabi industry as economically valuable plants (Keskin and Bunge, A. sosnowskyi Grossh., A. velenovskyi Nabelek, A. Kaçar, 2013; Tian et al., 2017; Adesso et al., 2018). trifoliastrum Hub.-Mor. & Matthews, A. uraniolimneus Karyological investigations carried out on taxa of the Boiss., A. gueruenensis Podlech, A. ciloensis Podlech, A. genus Astragalus showed that the diploid chromosome vaginans DC., A. szowitsii Fisch. & Mey., A. ermineus * Correspondence: [email protected] 232

This work is licensed under a Creative Commons Attribution 4.0 International License. MARTİN et al. / Turk J Bot

Matthews, A. surugensis Boiss. & Hausskn., A. zederbaueri Among the taxa in the sect. Hymenostegis, four Stadlmann, A. anthylloides Lam., A. halicacabus Lam., A. different somatic chromosomes (2n = 16, 18, 48, and 64) chardinii Boiss., and A. wagneri Bunge. are detected. The smallest chromosome size is found in A. ciloensis, with 1.37 µm, and the largest chromosome length 2. Materials and methods is found as 5.13 µm, for A. hymenocystis. The smallest Karyotypes were obtained from a number of members value for total haploid chromosome length is 21.10 µm, of Astragalus collected from different localities in Turkey in A. sosnowskyi, and the highest value is 69.40 µm, in A. (Table 1; Figure 1). lagopoides. The collected specimens were deposited in Aksaray Among the taxa in the sect. Anthylloidei, two different University Herbarium (AKSU). The seeds were germinated somatic chromosomes (2n = 16 and 48) are detected. The at room temperature. Then the root tips were pretreated in smallest and largest chromosome sizes are found in A. α-monobromonaphthalene at 4 °C for 16 h, fixed in fixative wagneri, with 1.59 µm and 4.18 µm. The smallest value (3 alcohol:1 acetic acid) for 24 h, hydrolyzed in 1 N HCl for for total haploid chromosome length is 19.27 µm, in 12 min, stained in 2% aceto-orcein at room temperature, A. anthylloides, and the highest value is 62.97 µm, in A. and squashed in 45% acetic acid. Permanent slides were surugensis. made by mounting in Depex. The 10 metaphase plates In the cytogenetic study, which was carried out on the from each species were used to obtain chromosomal data taxa belonging to the four different sections, only a pair of using an Olympus microscope and the chromosomal data satellite chromosomes is observed in A. uraniolimneus and were measured with software image analyses (Bs200ProP). a secondary constriction in A. velenovskyi. In Figure 4, very The chromosome classifications used the nomenclature weak correlations are calculated for intrachromosomal described by Levan et al. (1964), and the total haploid asymmetry parameters against interchromosomal lengths were measured based on Peruzzi et al. (2009). asymmetry parameters such as M -CV (r = 0.083). The intrachromosomal asymmetry, interchromosomal CA CL asymmetry, and intrachromosomal heterogeneity were 3.1. Sect. Macrophyllium 3.1.1. Astragalus oleaefolius determined with the parameters of MCA (Peruzzi and The diploid number is 2n = 16 and the karyotype formula Eroğlu, 2013), CVCL, and CVCI (Paszko, 2006), respectively. is 16m. The lengths of the chromosomes are between Finally, a scatter diagram was drawn between MCA and 1.88 and 2.89 µm. The total haploid and mean haploid CVCL. lengths are 19.38 and 2.42 µm, respectively. The values 3. Results of intrachromosomal asymmetry and interchromosomal The somatic metaphase chromosomes are given in asymmetry are 11.60 and 14.29, respectively. Figure 2, the monoploid idiograms in Figure 3, and the 3.1.2. Astragalus longifolius measurement data of chromosomes and the asymmetry The diploid number is 2n = 32 and the karyotype formula index values in Table 2 for Astragalus species. The following is 32m. The lengths of the chromosomes are between somatic chromosome numbers were found: 2n = 16 for A. 1.45 and 3.36 µm. The total haploid and mean haploid oleaefolius, A. dipodurus, A. cephalotes, and A. yukselii; 2n lengths are 37.46 and 2.34 µm, respectively. The values = 32 for A. longifolius; and 2n = 48 for A. isauricus in the of intrachromosomal asymmetry and interchromosomal sect. Macrophyllium; 2n = 16 for A. zohrabi, A. sosnowskyi, asymmetry are 11.86 and 20.33, respectively. A. velenovskyi, A. trifoliastrum, and A. uraniolimneus; 2n 3.1.3. Astragalus isauricus = 18 for A. hymenocystis; 2n = 48 for A. lagopoides, A. The diploid number is 2n = 48 and the karyotype formula gueruenensis, and A. ciloensis; and 2n = 64 for A. hirticalyx is 48m. The lengths of the chromosomes are between in the sect. Hymenostegis; 2n = 48 for A. vaginans in the sect. Hymenocoleus; 2n = 16 for A. szowitsii, A. ermineus, 1.51 and 3.64 µm. The total haploid and mean haploid A. zederbaueri, A. anthylloides, A. halicacabus, A. chardinii, lengths are 62.56 and 2.60 µm, respectively. The values and A. wagneri; and 2n = 48 for A. surugensis in the sect. of intrachromosomal asymmetry and interchromosomal Anthylloidei. asymmetry are 11.93 and 17.28, respectively. Among the taxa in the sect. Macrophyllium, three 3.1.4. Astragalus dipodurus different somatic chromosomes (2n = 16, 32, and 48) are The diploid number is 2n = 16 and the karyotype formula detected. The smallest chromosome size among the taxa is is 16m. The lengths of the chromosomes are between 1.45 µm, in A. longifolius. The largest chromosome size is 1.79 and 3.01 µm. The total haploid and mean haploid detected in A. isauricus, with 3.64 µm. The smallest value lengths are 18.94 and 2.36 µm, respectively. The values for total haploid chromosome length is 16.00 µm, in A. of intrachromosomal asymmetry and interchromosomal cephalotes, and the highest value is 62.56 µm, in A. isauricus. asymmetry are 9.28 and 16.13, respectively.

233 MARTİN et al. / Turk J Bot

Table 1. Localities and collector numbers of studied Astragalus.

Section Taxa Localities Voucher

B5 Aksaray: Taptuk Emre village, garden margins, 1406 m, A. oleaefolius S. Karaman 2542 07.06.2011 B9 Van: Erciş, between Hasanabdal and İşbaşı villages, moving A. longifolius S. Karaman 2678 slopes, 1855 m, 10.08.2011 C4 Konya: between Hadim and Taşkent, 3–5 A. isauricus S. Karaman 2690 Macrophyllium km from Hadim, steppe, 24.08.2011 A. dipodurus C4 Konya: Karasınır, steppe, field margins, 1100 m, 25.08.2011 S. Karaman 2693 A8 Artvin: Şavşat, Yalnızçam pass, naked slopes, 2630 m, A. cephalotes S. Karaman 2793 22.07.2013 C4 Konya: Hadim, 3 km from Hadim to Taşkent, steppe, A. yukselii S. Karaman 2691 1580 m, 24.08.2011 A. hymenocystis B9 Van: Gürpınar, Güzeldere pass, steppe, 2760 m, 20.07.2013 S. Karaman 2752 A. lagopoides B9 Van: Erciş, Pay village, steppe, 2050 m, 14.06.2012 S. Karaman 2716 B9 Van: Hoşap–Başkale road, Güzeldere pass, steppe, 2616 m, A. hirticalyx S. Karaman 2681 11.08.2011 A. zohrabi B9 Van: Erciş, Deliçay, Pay village, steppe, 2095 m, 10.08.2011 S. Karaman 2676 A. sosnowskyi A8 Erzurum: Dutlu Mountain, NW Oltu, 2300 m, 15.07.2011 S. Karaman 2666 B9 Van: Erciş, between Deliçay and Pay villages, steppe, 2000 m, A.velenovskyi S. Karaman 2759 Hymenostegis 23.06.2013 B9 Van: Gürpınar, between Hamurkesen and Işıkpınar villages, A. trifoliastrum S. Karaman 2651 steppe, 2155 m, 13.07.2011 A8 Artvin: Yusufeli, Ziyaret Mountain, Narlık plateau, alpine A. uraniolimneus S. Karaman 2799 meadows, 2630 m, 03.09.2011 B6 Sivas: 46 km from Gürün to Pınarbaşı, roadsides and steppe, A. gueruenensis S. Karaman 2800 1700 m, 07.10.2013 A. ciloensis B9 Van: Hoşap, Çuh Gediği, steppe, 2700 m, 11.08.2011 S. Karaman 2679 C6 Kahramanmaraş: Çağlayancerit, Ziyaret Ağaçlı hill, steppe, Hymenocoleus A. vaginans S. Karaman 2672 1200–1300 m, 22.07.2011 A. szowitsii B10 Ağrı: Doğubeyazıt, Iran border, steppe, 1600 m, 25.06.2013 S. Karaman 2771 B9 Van: Bahçesaray, Karapit pass, rock slits, 2970–3000 m, A. ermineus S. Karaman 2674 09.08.2011 C7 Şanlıurfa: 24 km from Şanlıurfa to Hilvan, calcareous slopes, A. surugensis S. Karaman 2697 700 m, 07.05.2012 C4 Konya: Surroundings of Altınapa dam, calcareous slopes, A. zederbaueri S. Karaman 2624 1290 m, 07.07.2011 Anthylloidei A4 Ankara: Ayaş, Aysantı pass, calcareous slopes, S. Karaman 2617 & A. anthylloides 1160 m, 28.06.2011 D. Bozer A. halicacabus B9 Van: Muradiye, Hangediği, steppe, 2268 m, 12.07.2011 S. Karaman 2634 B8 Erzurum: 27 km from Horasan to Ağrı, steppe, 1970 m, A. chardinii S. Karaman 2656 14.07.2011 B10 Ağrı: Doğubeyazıt, slopes of İshakpaşa palace, steppe, A. wagneri S. Karaman 2655 1994 m, 14.07.2011

234 MARTİN et al. / Turk J Bot

Figure 1. Habitat of Astragalus sections Macrophyllium, Hymenostegis, Hymenocoleus, and Anthylloidei: a A. oleaefolius, b A. longifolius, c A. isauricus, d A. dipodurus, e A. cephalotes, f A. yukselii, g A. hymenocystis, h A. lagopoides, i A. hirticalyx, j A. zohrabi, k A. sosnowskyi, l A. velenovskyi, m A. trifoliastrum, n A. uraniolimneus, o A. gueruenensis, p A. ciloensis, q A. vaginans, r A. szowitsii, s A. ermineus, t A. surugensis, u A. zederbaueri, v A. anthylloides, w A. halicacabus, x A. chardinii, y A. wagneri.

235 MARTİN et al. / Turk J Bot

Figure 2. Somatic metaphase chromosomes of Astragalus sections Macrophyllium, Hymenostegis, Hymenocoleus, and Anthylloidei: a A. oleaefolius, b A. longifolius, c A. isauricus, d A. dipodurus, e A. cephalotes, f A. yukselii, g A. hymenocystis, h A. lagopoides, i A. hirticalyx, j A. zohrabi, k A. sosnowskyi, l A. velenovskyi, m A. trifoliastrum, n A. uraniolimneus, o A. gueruenensis, p A. ciloensis, q A. vaginans, r A. szowitsii, s A. ermineus, t A. surugensis, u A. zederbaueri, v A. anthylloides, w A. halicacabus, x A. chardinii, y A. wagneri. Long arrow, secondary constriction and short arrow, satellites (scale bar = 10 µm).

236 MARTİN et al. / Turk J Bot

Figure 3. Idiograms of Astragalus sections Macrophyllium, Hymenostegis, Hymenocoleus, and Anthylloidei: a A. oleaefolius, b A. longifolius, c A. isauricus, d A. dipodurus, e A. cephalotes, f A. yukselii, g A. hymenocystis, h A. lagopoides, i A. hirticalyx, j A. zohrabi, k A. sosnowskyi, l A. velenovskyi with a secondary constriction, m A. trifoliastrum, n A. uraniolimneus with satellite, o A. gueruenensis, p A. ciloensis, q A. vaginans, r A. szowitsii, s A. ermineus, t A. surugensis, u A. zederbaueri, v A. anthylloides, w A. halicacabus, x A. chardinii, y A. wagneri.

237 MARTİN et al. / Turk J Bot

3.1.5. Astragalus cephalotes 3.2.6. Astragalus velenovskyi The diploid number is 2n = 16 and the karyotype formula The diploid number is 2n = 16 and the karyotype formula is 16m. The lengths of the chromosomes are between is 16m. The lengths of the chromosomes are between 1.61 and 2.67 µm. The total haploid and mean haploid 2.47 and 4.45 µm. The total haploid and mean haploid lengths are 16.00 and 2.00 µm, respectively. The values lengths are 27.49 and 3.44 µm, respectively. The values of intrachromosomal asymmetry and interchromosomal of intrachromosomal asymmetry and interchromosomal asymmetry are 13.00 and 17.51, respectively. asymmetry are 12.86 and 19.80, respectively. 3.1.6. Astragalus yukselii 3.2.7. Astragalus trifoliastrum The diploid number is 2n = 16 and the karyotype formula The diploid number is 2n = 16 and the karyotype formula is is 16m. The lengths of the chromosomes are between 14m + 2sm. The lengths of the chromosomes are between 1.59 and 2.75 µm. The total haploid and mean haploid 2.27 and 4.12 µm. The total haploid and mean haploid lengths are 17.62 and 2.20 µm, respectively. The values lengths are 25.17 and 3.15 µm, respectively. The values of intrachromosomal asymmetry and interchromosomal of intrachromosomal asymmetry and interchromosomal asymmetry are 12.56 and 16.76, respectively. asymmetry are 11.24 and 18.95, respectively. 3.2. Sect. Hymenostegis 3.2.8. Astragalus uraniolimneus The diploid number is 2n = 16 and the karyotype formula 3.2.1. Astragalus hymenocystis is 16m (2sat). The lengths of the chromosomes are between The diploid number is 2n = 18 and the karyotype formula 2.42 and 3.64 µm. The total haploid and mean haploid is 18m. The lengths of the chromosomes are between lengths are 25.20 and 3.15 µm, respectively. The values 2.59 and 5.13 µm. The total haploid and mean haploid of intrachromosomal asymmetry and interchromosomal lengths are 33.44 and 3.71 µm, respectively. The values asymmetry are 10.22 and 13.25, respectively. of intrachromosomal asymmetry and interchromosomal 3.2.9. Astragalus gueruenensis asymmetry are 10.53 and 20.40, respectively. The diploid number is 2n = 48 and the karyotype formula 3.2.2. Astragalus lagopoides is 48m. The lengths of the chromosomes are between The diploid number is 2n = 48 and the karyotype formula 1.56 and 3.28 µm. The total haploid and mean haploid is 48m. The lengths of the chromosomes are between lengths are 54.59 and 2.27 µm, respectively. The values 1.81 and 4.25 µm. The total haploid and mean haploid of intrachromosomal asymmetry and interchromosomal lengths are 69.40 and 2.89 µm, respectively. The values asymmetry are 10.54 and 17.97, respectively. of intrachromosomal asymmetry and interchromosomal 3.2.10. Astragalus ciloensis asymmetry are 11.09 and 21.14, respectively. The diploid number is 2n = 48 and the karyotype formula 3.2.3. Astragalus hirticalyx is 48m. The lengths of the chromosomes are between The diploid number is 2n = 64 and the karyotype formula 1.37 and 3.40 µm. The total haploid and mean haploid is 64m. The lengths of the chromosomes are between lengths are 56.21 and 2.34 µm, respectively. The values 1.67 and 4.44 µm. The total haploid and mean haploid of intrachromosomal asymmetry and interchromosomal lengths are 85.67 and 2.68 µm, respectively. The values asymmetry are 10.28 and 22.65, respectively. of intrachromosomal asymmetry and interchromosomal 3.3. Sect. Hymenocoleus asymmetry are 9.86 and 20.81, respectively. 3.3.1. Astragalus vaginans 3.2.4. Astragalus zohrabi The diploid number is 2n = 48 and the karyotype formula The diploid number is 2n = 16 and the karyotype formula is is 48m. The lengths of the chromosomes are between 14m + 2sm. The lengths of the chromosomes are between 1.45 and 3.27 µm. The total haploid and mean haploid 2.16 and 4.02 µm. The total haploid and mean haploid lengths are 57.08 and 2.38 µm, respectively. The values lengths are 23.47 and 2.93 µm, respectively. The values of intrachromosomal asymmetry and interchromosomal of intrachromosomal asymmetry and interchromosomal asymmetry are 11.76 and 18.47, respectively. asymmetry are 15.48 and 20.62, respectively. 3.4. Sect. Anthylloidei 3.2.5. Astragalus sosnowskyi 3.4.1. Astragalus szowitsii The diploid number is 2n = 16 and the karyotype formula The diploid number is 2n = 16 and the karyotype formula is 16m. The lengths of the chromosomes are between is 16m. The lengths of the chromosomes are between 2.01 and 3.24 µm. The total haploid and mean haploid 2.92 and 4.09 µm. The total haploid and mean haploid lengths are 21.10 and 2.63 µm, respectively. The values lengths are 27.51 and 3.44 µm, respectively. The values of intrachromosomal asymmetry and interchromosomal of intrachromosomal asymmetry and interchromosomal asymmetry are 9.70 and 15.32, respectively. asymmetry are 7.22 and 11.26, respectively.

238 MARTİN et al. / Turk J Bot

Table 2. The measurement data of chromosomes of Astragalus taxa. SC, shortest chromosome length; LC, longest chromosome length; THL, total haploid chromosome length; MHL, mean haploid chromosome length; CN, chromosome number; KF, karyotype formula;

CVCI, intrachromosomal heterogeneity; CVCL, coefficient of variation of chromosome length;CA M , mean centromeric asymmetry.

Taxa SC (µm) LC (µm) THL (µm) MHL (µm) CN and KF CVCI CVCL MCA A. oleaefolius 1.88 2.89 19.38 2.42 2n = 16 = 16m 3.45 14.29 11.60 A. longifolius 1.45 3.36 37.46 2.34 2n = 32 = 32m 5.61 20.33 11.86 A. isauricus 1.51 3.64 62.56 2.60 2n = 48 = 48m 5.53 17.28 11.93 A. dipodurus 1.79 3.01 18.94 2.36 2n = 16 = 16m 3.48 16.13 9.28 A. cephalotes 1.61 2.67 16.00 2.00 2n = 16 = 16m 6.44 17.51 13.00 A. yukselii 1.59 2.75 17.62 2.20 2n = 16 = 16m 6.04 16.76 12.56 A. hymenocystis 2.59 5.13 33.44 3.71 2n = 18 = 18m 6.50 20.40 10.53 A. lagopoides 1.81 4.25 69.40 2.89 2n = 48 = 48m 3.99 21.14 11.09 A. hirticalyx 1.67 4.44 85.67 2.68 2n = 64 = 64m 3.90 20.81 9.86 A. zohrabi 2.16 4.02 23.47 2.93 2n = 16 = 14m + 2sm 15.94 20.62 15.48 A. sosnowskyi 2.01 3.24 21.10 2.63 2n = 16 = 16m 4.20 15.32 9.70 A. velenovskyi 2.47 4.45 27.49 3.44 2n = 16 = 16m 4.64 19.80 12.86 A. trifoliastrum 2.27 4.12 25.17 3.15 2n = 16 = 14m + 2sm 8.41 18.95 11.24 A. uraniolimneus 2.42 3.64 25.20 3.15 2n = 16 = 16m (2sat) 3.72 13.25 10.22 A. gueruenensis 1.56 3.28 54.59 2.27 2n = 48 = 48m 5.34 17.97 10.54 A. ciloensis 1.37 3.40 56.21 2.34 2n = 48 = 48m 5.57 22.65 10.28 A. vaginans 1.45 3.27 57.08 2.38 2n = 48 = 48m 4.47 18.47 11.76 A. szowitsii 2.92 4.09 27.51 3.44 2n = 16 = 16m 2.27 11.26 7.22 A. ermineus 2.04 3.70 23.23 2.90 2n = 16 = 16m 4.74 18.83 8.25 A. surugensis 1.88 3.43 62.97 2.62 2n = 48 = 2M + 46m 6.14 16.79 10.78 A. zederbaueri 2.00 3.82 23.52 2.94 2n = 16 = 16m 2.52 21.19 7.91 A. anthylloides 1.85 3.13 19.27 2.40 2n = 16 = 16m 6.38 16.81 12.81 A. halicacabus 1.88 3.49 21.29 2.66 2n = 16 = 16m 4.68 20.32 11.40 A. chardinii 1.89 3.12 19.37 2.42 2n = 16 = 16m 4.46 17.28 10.00 A. wagneri 1.59 4.18 23.05 2.88 2n = 16 = 16m 3.18 28.25 9.29

3.4.2. Astragalus ermineus of intrachromosomal asymmetry and interchromosomal The diploid number is 2n = 16 and the karyotype formula asymmetry are 7.91 and 21.19, respectively. is 16m. The lengths of the chromosomes are between 3.4.5. Astragalus anthylloides 2.04 and 3.70 µm. The total haploid and mean haploid The diploid number is 2n = 16 and the karyotype formula lengths are 23.23 and 2.90 µm, respectively. The values is 16m. The lengths of the chromosomes are between of intrachromosomal asymmetry and interchromosomal 1.85 and 3.13 µm. The total haploid and mean haploid asymmetry are 8.25 and 18.83, respectively. lengths are 19.27 and 2.40 µm, respectively. The values 3.4.3. Astragalus surugensis of intrachromosomal asymmetry and interchromosomal The diploid number is 2n = 48 and the karyotype formula asymmetry are 12.81 and 16.81, respectively. is 2M + 46m. The lengths of the chromosomes are between 3.4.6. Astragalus halicacabus 1.88 and 3.43 µm. The total haploid and mean haploid The diploid number is 2n = 16 and the karyotype formula lengths are 62.97 and 2.62 µm, respectively. The values is 16m. The lengths of the chromosomes are between of intrachromosomal asymmetry and interchromosomal 1.88 and 3.49 µm. The total haploid and mean haploid asymmetry are 10.78 and 16.79, respectively. lengths are 21.29 and 2.66 µm, respectively. The values 3.4.4. Astragalus zederbaueri of intrachromosomal asymmetry and interchromosomal The diploid number is 2n = 16 and the karyotype formula asymmetry are 11.40 and 20.32, respectively. is 16m. The lengths of the chromosomes are between 3.4.7. Astragalus chardinii 2.00 and 3.82 µm. The total haploid and mean haploid The diploid number is 2n = 16 and the karyotype formula lengths are 23.52 and 2.94 µm, respectively. The values is 16m. The lengths of the chromosomes are between

239 MARTİN et al. / Turk J Bot

Figure 4. Scatter diagram between MCA and CVCL: Sections of Macrophyllium, Hymenostegis, Hymenocoleus, and Anthylloidei. a A. oleaefolius, b A. longifolius, c A. isauricus, d A. dipodurus, e A. cephalotes, f A. yukselii, g A. hymenocystis, h A. lagopoides, i A. hirticalyx, j A. zohrabi, k A. sosnowskyi, l A. velenovskyi, m A. trifoliastrum, n A. uraniolimneus, o A. gueruenensis, p A. ciloensis, q A. vaginans, r A. szowitsii, s A. ermineus, t A. surugensis, u A. zederbaueri, v A. anthylloides, w A. halicacabus, x A. chardinii, y A. wagneri.

1.89 and 3.12 µm. The total haploid and mean haploid karyological data suggested that three Astragalus taxa lengths are 19.37 and 2.42 µm, respectively. The values had median and submedian chromosomes except for A. of intrachromosomal asymmetry and interchromosomal cariensis Boiss. (Martin et al., 2008). asymmetry are 10.00 and 17.28, respectively. In the present study, satellites were found in the first 3.4.8. Astragalus wagneri pair in A. uraniolimneus (Figure 2n, short arrow). Previous The diploid number is 2n = 16 and the karyotype formula studies reported that A. chlorostachys Lindl. had three is 16m. The lengths of the chromosomes are between satellite chromosome pairs (Manandhar and Sakya, 2004), 1.59 and 4.18 µm. The total haploid and mean haploid A. stella L. had a single satellite chromosome pair (Badr lengths are 23.05 and 2.88 µm, respectively. The values and Sharawy, 2007), and A. nezaketae A.Duran & Aytaç of intrachromosomal asymmetry and interchromosomal had two satellite chromosome pairs (Martin et al., 2008). asymmetry are 9.29 and 28.25, respectively. In addition, more satellites are likely to be determined. However, the satellites are not always detectable, depending 4. Discussion on the chromatin condensation (Baziz et al., 2014). This study reports the diploid chromosome numbers In the present study, a secondary constriction was and karyotypes of 25 species of Astragalus from the detected in A. velenovskyi (Figure 2l, long arrow). sections Macrophyllium, Hymenostegis, Hymenocoleus, Previous studies reported that secondary constrictions and Anthylloidei. In all taxa, the karyotype results were were determined in A. cruciatus Link., A. armatus obtained for the first time. The cytological data confirmed subsp. tragacanthoides (Desf.) Maire, A. armatus subsp. that the chromosome numbers of Astragalus taxa are 2n = numidicus (Coss. & Dur.) Maire (Baziz et al., 2014), A. 16, 18, 32, 48, and 64 with the basic numbers of x = 8 and 9. mongholicus Bunge, and A. membranaceus (Fisch.) Bunge All Astragalus taxa examined in this study show a (Ma et al., 1984). similar karyomorphology including only median and Karyotype asymmetry is one of the most important submedian chromosomes. Recent studies also confirm parameters in chromosomal data (Eroğlu et al., similar karyotypes for Astragalus. Konichenko and 2013). MCA and CVCL are the most reliable parameters Selyutina (2017) reported the karyotype formula of A. among asymmetry indexes (Peruzzi and Eroğlu, chorinensis Bunge as 2n = 2m + 14sm. The population of 2013). Intrachromosomal asymmetry is characterized A. sericeocanus Gontsch. was divided into three karyotype by a karyotype with a predominance of terminal/ groups with 2n = 16 = 4m + 12sm, 2n = 16 = 6m + 10sm, subterminal chromosomes and is calculated with MCA. and 2n = 16 = 8m + 8sm (Konichenko et al., 2014). The Interchromosomal asymmetry is characterized by highly

240 MARTİN et al. / Turk J Bot

heterogeneous chromosome sizes and is calculated There is a weak correlation between values of CAM with CVCL (Peruzzi and Eroğlu, 2013). All Astragalus and CVCL (r = 0.083) (Figure 4). The weak correlation taxa are characterized by symmetrical karyotypes with is likely due to the CVCL parameter, which gives reliable a predominance of median/submedian chromosomes data with the MCA parameter (Peruzzi and Eroğlu, as a common condition in the genus (Manandhar and 2013). Sakya, 2004; Badr and Sharawy, 2007; Martin et al., 2008; In Figure 4, the four sections of the genus Astragalus Baziz et al., 2014). In contrast, Baziz et al. (2014) reported have different karyotypes in terms of asymmetry that A. pseudotrigonus Batt. & Trab. and A. cruciatus had degrees: the sections Macrophyllium and Hymenocoleus asymmetrical karyotypes. with relatively average intrachromosomal and In the sect. Hymenostegis, the most symmetrical interchromosomal asymmetry, the sect. Hymenostegis karyotype is A. sosnowskyi and the most asymmetrical is with higher intrachromosomal asymmetry, and the

A. zohrabi based on MCA. The most symmetrical one is A. sect. Anthylloidei with higher interchromosomal uraniolimneus and the most asymmetrical one is A. ciloensis asymmetry. in CVCL. In the present study, new chromosomal data In the sect. Anthylloidei, the most symmetrical are reported for 25 species of the genus Astragalus. karyotype is A. szowitsii based on all index values. However, Karyotype analyses were carried out in detail for the the asymmetrical karyotypes are different. While the first time for all taxa. More karyological data are needed most asymmetrical one is A. anthylloides in MCA, the most to contribute to the cytotaxonomy of Astragalus. asymmetrical one is A. wagneri in CVCL. In all sections, the most symmetrical karyotype Acknowledgments is A. szowitsii based on all index values. However, We wish to thank TÜBİTAK (project number: TBAG- the asymmetrical karyotypes are different. While the 110T911 and KBAG-113Z899) for its financial support most asymmetrical one is A. zohrabi in MCA, the most and we are grateful to Dr Z Aytaç and Dr M Ekici (Gazi asymmetrical one is A. wagneri in CVCL. University) for their invaluable advice.

References

Adesso S, Russo R, Quaroni A, Autore G, Marzocco S (2018). Hung LY (1984). Karyotype analysis of 5 species of genus Astragalus. Astragalus membranaceus extract attenuates inflammation and Acta Phytotax Sin 22: 125-127. oxidative stress in intestinal epithelial cells via NF- B activation K İlçim A, Behçet L (2016). Astragalus topalanense (Fabaceae), a new and Nrf2 response. Int J Mol Sci 19: 800-814. species from Turkey. Turk J Bot 40: 74-80. Aytaç Z (1997). The revision of the section Dasyphyllium Bunge of Kandemir N, Korkmaz H, Engin A (1996). The morphological and the genus Astragalus L. of Turkey. Turk J Bot 21: 31-59. anatomical properties of Astragalus barba-jovis DC. var. barba- Badr A, Sharawy SM (2007). Karyotype analysis and systematic jovis (Fabaceae). Turk J Bot 20: 291-299. relationships in the Egyptian Astragalus L. (Fabaceae). Int J Bot Karaman Erkul S, Aytaç Z, Ekici M (2016). Synopsis of the sect. 3: 147-159. Hymenocoleus, sect. Hymenostegis, and sect. Macrophyllium Baziz K, Benamara-Bellagha M, Pustahija F, Brown SC, Siljak- belonging to Astragalus (Fabaceae) in Turkey. Turk J Bot 40: Yakovlev S, Khalfallah N (2014). First karyotype analysis, 412-418. physical rDNA mapping, and genome size assessment in 4 North African Astragalus taxa (Fabaceae). Turk J Bot 38: 1248- Kazem Y, Houshmand S, Dadane GZ (2010). Karyotype analysis of 1258. Astragalus effusus Bunge (Fabaceae). Caryologia 63: 257-261. Borjian A, Assadi M, Nejadsattari T, Maassoumi AA (2012). Based Keskin C, Kaçar S (2013). Fatty acid composition of root and shoot on cytotaxonomical and morphological evidence Astragalus samples of some Astragalus L. (Fabaceae) taxa growing in the remotiflorus subsp. nigrohirsutus is raised to species rank. Iran east and southeast of Turkey. Turk J Biol 37: 122-128. J Bot 18: 98-103. Konichenko ES, Selyutina IY (2017). The karyotype of some rare Davina JR, Gomez-Sosa E (1993). Cariotipo de siete especies del species of Astragalus and Oxytropis (Fabaceae). Turczaninowia genero Astragalus (Leguminosae) de la Argentina. Bol Soc 20: 31-38. Argent Bot 29: 197-201. Konichenko ES, Selyutina IY, Dorogina OV, Sandanov D (2014). Eroğlu HE, Şimşek N, Koç M, Hamzaoğlu E (2013). Karyotype Karyotype studies endemic plant species Astragalus analysis of some Minuartia L. (Caryophyllaceae) taxa. Plant sericeocanus Gonsch. (Fabaceae) around Like Baikal, Siberia. Syst Evol 299: 67-73. Caryologia 67: 172-177.

241 MARTİN et al. / Turk J Bot

Levan AK, Fredga K, Sandberg AA (1964). Nomenclature for Peruzzi L, Leitch IJ, Caparelli KF (2009). Chromosome diversity and centromeric position on chromosomes. Hereditas 52: 201-220. evolution in Liliaceae. Ann Bot 103: 459-475. Liston A (1990). Chromosome counts in Astragalus (Fabaceae). Peruzzi L, Eroğlu HE (2013). Karyotype asymmetry: again, how to Madroño 37: 59-60. measure and what to measure? Comp Cytogen 7: 1-9. Ma XH, Qin RL, Xing WB (1984). Chromosome observations of Rani S, Kumari S, Gupta RC, Chahota RK (2014). Cytological studies some medical plants in Xinjiang. Acta Phytotaxon Sin 22: 243- of Angiosperms (174 species) from District Kangra, Himachal 249. Pradesh (India). Plant Syst Evol 300: 851-862. Maassoumi AA (1987). Notes on the genus Astragalus in Iran I, Sheidai M, Shahin Z, Jalal I (2009). New chromosome number cytotaxonomic studies on some species. Iran J Bot 3: 117-128. reports in tragacanthic Astragalus species. Caryologia 62: 30- 36. Magulaev AJ (1989). Chromosome numbers in some Astragalus (Fabaceae) species of the Caucasus flora. Bot Žhurn (Moscow Spellenberg R (1981). Chromosome numbers and their & Leningrad) 74: 1519-1521. cytotaxonomic significance for North American Astragalus (Fabaceae). Taxon 25: 463-472. Manandhar L, Sakya SR (2004). Cytotaxonomic studies in two species of Astragalus. J Cytol Genet 5: 13-20. Tian Z, Liu Y, Yang B, Zhang J, He H, Ge H, Wu Y, Shen Z (2017). Astagalus polysaccharide attenuates murine colitis through Martin E, Duran A, Dinç M, Erişen S, Babaoğlu M (2008). Karyotype inhibiton of the NLRP3 inflammasome. Planta Med 83: 70-77. analyses of four Astragalus L. (Fabaceae) species from Turkey. Phytologia 90: 147-159. Ward DE, Spellenberg R (1988). Chromosome counts of angiosperms from New Mexico and adjacent areas. Phytologia 64: 390-398. Paszko B (2006). A critical review and a new proposal of karyotype asymmetry indices. Plant Syst Evol 258: 39-48.

242