Karyotype Investigations in Iranian Scorzonera Tortuosissima and S

Karyotype Investigations in Iranian Scorzonera tortuosissima and S. intricata, Asteraceae

Firouzeh Bordbar*, Parinaz Makki and Fereshteh Alifarsangi

Department of Biology, Faculty of Sciences, Shahid Bahonar University of Kerman, Kerman, Iran

Received September 9, 2018; accepted October 31, 2018

Summary Somatic chromosomes of two Scorzonera species from subgenus Scorzonera, section Intricatae collected from four localities in southeast Iran were observed. In S. tortuosissima, 2n=28 was observed. S. intricata revealed two chromosome numbers of 2n=20 and 21. The chromosome numbers of 2n=21 had one B chromosome then the basic number is x=10 which is a new basic chromosome number for Scorzonera. Both species showed a predominance of metacentric chromosomes in each karyotype. The karyotype was formulated as 12m+2sm for S. tortuosissima and 8m+2sm for S. intricata.

Key words Asteraceae, B chromosome, Chromosome number, Iran, Karyotype, Scorzonera tortuosissima, Scorzonera intricata.

The Scorzonera L. is one of the large taxa of Astera- are deposited in Herbarium Mirtadzadini (MIR). ceae, with about 180 species and is belonged to the sub- Karyotype studies were made on somatic metaphase tribe Scorzonerinae Dumort. of the tribe Cichorieae. It is cells of the growing roots from seeds in petri dishes widely spread among the arid regions of Eurasia, Central using a standard squash technique according to Bord- Asia and northern Africa (Bremer and Anderberg 1994, bar et al. (2017). Root tips of 1–2 cm were cut off and Nazarova 1997). This genus is known as a taxonomi- pretreated with saturated α-monobromonaphthalene at cally difficult group with several complexes of closely 4°C for 8 h, then fixed in a fresh Carnoy’s fixative related taxa (Nazarova 1997). (3 : 1=ethanol : glacial acetic acid) for 24 h and stored at Chromosomal investigations have proved to be useful 4°C until use. Root tips were washed in distilled water to for tracing karyotype evolution and clarifying phyloge- remove the fixative, hydrolyzed in 1 M HCl for 5–7 min netic relationships among taxa (Soltis 2014). One of the at 60°C and stained in aceto-iron hematoxylin for 24 h most prominent karyotype investigations in Scorzonera at 30°C. Squash was done gently in a drop of 45% acetic was done by Nazarova (1997) who reported karyotypes acid on a glass slide. The best mitotic metaphases cells of 22 species and also reviewed previous chromosome were photographed using an Olympus BH-2 microscope reports. By the karyotype analysis, she supported the equipped with a digital camera. For each species lengths sectional delimitation in Scorzonera and also indicated of both arms of each chromosome in the complement the relationships between this genus and closely related in five cells were measured. Karyological parameters taxa. Therefore the karyotype investigations in this ge- of total chromosome length of the haploid complement nus are of particular importance. (HCL), total form percentage (TF%, Huziwara 1962), In the present work, we studied the karyotype in two intrachromosomal asymmetry index (A1, Romero-Zarco Scorzonera species, distributed in Iran in order to de- 1986), interchromosomal asymmetry index (A2, Rome- posit karyotype data for the taxonomic and phylogenetic ro-Zarco 1986), Arano index of karyotype asymmetry study of Iranian Scorzonera. (AsK%, Arano 1963), karyotype formula and classification of Stebbins (1971) were estimated and their ideo- Materials and methods grams were obtained using IdeoKar 1.1 software (Mirza- ghaderi and Marzangi 2015). The cytological investigations were performed on the specimens of Scorzonera tortuosissima Boiss. and S. in- Results tricata Boiss. belonging to subgenus Scorzonera, section Intricatae (Boiss.) Lipsch. (Rechinger 1977) collected Chromosome number and karyotype of S. tortuosis- from Iran. For both species, we studied 20–25 individu- sima was examined in 20 individuals of two localities als of two different localities (Table 1). The specimens (Table 1). The chromosome number of 2n=28 (Fig. 1A, D) was observed in all individuals. The karyotype was * Corresponding author, e-mail: [email protected] formulated as 2n=12m+2sm and the class of Stebbins DOI: 10.1508/cytologia.84.69 (1971) was 1B (Table 2). Secondary constriction was in- 70 F. Bordbar et al. Cytologia 84(1)

Table 1. Name of the plant species, locality information, chromosome numbers and the number of individuals.

Chromosome Number of Taxon Locality information number individuals

S. tortuosissima Iran, SE, Kerman, Bardsir, 1 km to Bidkhun, 2550 m, N29°39′43.6″, E56°30′44.2″, 2n=28 10 19.VI.2015, Bordbar and Makki 3001 (MIR). Iran, SE, NE Kerman, Dehbala village, N30°17′09.07″, E57°19′09.06″, 2386 m, 11.VI.2015, 2n=28 10 Bordbar and Makki 3002 (MIR) S. intricata Iran, SE, Kerman, Bardsir, 2 km to Bidkhun, N29°38′53.4″, E56°30′48.8″, 2500 m, 2n=20 4 19.VI.2015, Bordbar and Makki 3004 (MIR) 2n=21 16 Iran, SE, Kerman, Negar to Baft, Sangesayad pass, 08.VII.2018, Bordbar and Mirtadzadini 2n=20 10 3005 (MIR)

Fig. 1. Metaphase chromosomes and Karyotypes of S. tortuosissima (A, D) and S. intricata (B, E, C, F). B letter in B and arrow in b denote B chromosome. S indicates chromosome pair bearing satellite. Scale bars=10 µm. dicated for two chromosome pairs (Figs. 1D, 2A). The chromosome number and karyotype of S. intricata were investigated on 25 individuals of two localities (Table 1). This species possessed two chromosome number of 2n=21 (Figs. 1B, E, 2B) and 2n=20 (Fig. 1C, F). Chromosome number of 2n=20 was found in both localities, the accession numbers 3004 (MIR) and 3005 (MIR) and 2n=21 existed in most individuals of accession number 3004 (MIR) (Table 1). Although this chromosome in all the observations morphologically looks like chromosomes (Figs. 1B, E, 2B), one chromosome Fig. 2. Haploid idiograms of S. tortuosissima (A) and S. intricata in the plants with 2n=21, considered as B chromosome (B). B letter denotes B chromosome. Scale bar=10 µm. because of irregular distribution and absence in some individuals of one population (Jones et al. 2008). The indicated for two pairs of chromosomes (Fig. 1E, F). chromosome number of this taxon is given here for the According to the classiﬁcation of Stebbins (1971), the ﬁrst time. The karyotypes were formulated as 8m+2sm karyotype of S. intricata presented symmetry type 2B and 8m+2sm+1B. Moreover secondary constriction was (Table 2). 2019 Karyotype Investigations in Iranian Scorzonera 71

Table 2. Karyotype features of S. tortuosissima and S. intricata.

Taxon HCL (µm) TF% A1 A2 AsK% Karyotype formulae Stebbins class S. tortousissima 52.86 42.04 0.26 0.33 57.96 12m+2sm 1B S. intricata 58.13 39.75 0.27 0.28 60.25 8m+2sm 2B 8m+2sm+1B

According to karyological features measured for S. metry. The high symmetry of karyotypes present by the tortuosissima and S. intricata; TF% (42.4 and 39.75, predominance of metacentric chromosomes in Scorzo- respectively), AsK% (57.96 and 60.25, respectively), A1 nera species. The review of the previous reports as well (0.26 and 0.27, respectively) and A2 (0.33 and 0.28, re- as the karyotype investigations in Scorzonera species spectively) and also Stebbins classification, both species from Armenia and middle Asia by Nazarova (1997) indicate almost symmetric karyotypes (Table 2). also confirmed the prevalence of metacentric chromosomes in this genus. Karyomorphological studies on Discussion Scorzonera species from flora of Turkey (Martïn et al. 2012, Altinordu et al. 2015) and Iberian Peninsula (Díaz Scorzonera is represented by 56 species in Iran of De La Guardia and Blanca 1987) also rarely exhibited which 28 species are endemic for the flora of Iran (Re- asymmetric karyotypes. chinger 1977, Safavi et al. 2013, Safavi 2016). In this However, and x=6 and 7 are stable basic numbers but study, we investigated the karyotype of two species S. the species belong to the genus have been insufficiently intricata and S. tortuosissima sect. Intricatae belonging studied. Therefore it seems that more extensive karyo- to the flora of Iran. S. tortuosissima is widely distributed logic and genomic data are needed to the assessment of in south, east and center of Iran as well as Afghanistan the karyotype evolution in Scorzonera. and Pakistan (Rechinger 1977, Safavi et al. 2013). S. intricata is an endemic species for the flora of Iran which Acknowledgements distributed in south-east, east and center of Iran (Rech- inger 1977, Safavi et al. 2013). This study was supported by a greatly appreciated Our result confirmed the previous count of 2n=28 grant from Shahid Bahonar University of Kerman. by Razaq et al. (1994) for S. tortuosissima and was in disagreement with the 2n=42 reported by Khatoon and References Ali (1993) for this species. The results of this study for S. intricata revealed x=10 and two different chromo- Altinordu, F., Martïn, E., Makbul, S., Coşkunçelebï, K. and Gültepe, some numbers of 2n=20 in accession numbers 3004 M. 2015. Cytogenetic studies on some Scorzonera L. s.l. (Astera- ceae) taxa from Turkey. Turk. J. Bot. 39: 429–438. (MIR) and 3005 (MIR) and 2n=21 in some individuals Arano, H. 1963. Cytological studies in subfamily Carduoideae (Com- of accession number 3004 (MIR). Therefore the plants positae) of Japan. IX. Bot. Mag. Tokyo 76: 32–39. of 2n=21 showed one accessory or B chromosome. The Bordbar, F., Baqeri, M. and Mirtadzadini, M. 2017. New chromosome occurrence of one chromosome has been informed pre- counts in Cousinia (Asteraceae) from flora of Iran. Cytologia 82: viously for few species of Scorzonera (Nazarova 1997). 485–488. Bremer, K. and Anderberg, A. A. 1994. Asteraceae: Cladistics and The chromosome number of 2n=21 has been previously Classification. Timbers Press, Portland. reported to occur only in S. tau-saghyz Lipsch. et Bosse. Díaz De La Guardia, C. and Blanca, G. 1987. Karyology of the Scor- from sect. Egregiae (Kult.) Lipsch., subgen. Scorzonera zonera (Compositae) species from the Iberian Peninsula. Plant (Krajevoy 1934). However not only polyploid cytotypes, Syst. Evol. 156: 29–42. 2n=14, 21, 28, 42 but also aneuploids with 2n=15, 16, Huziwara, Y. 1962. Karyotype analysis in some genera of Composi- 17, 18, 22 and 24 have been reported for S. tau-saghyz tae. VIII. Further studies on the chromosomes of Aster. Am. J. Bot. 49: 116–119. (Plotnikova 1933, Krajevoy 1934, Poddubnaja-Arnoldi Jones, R. N., Viegas, W. and Houben, A. 2008. A century of B chro- et al. 1934). mosomes in plants: So what? Ann. Bot. 101: 767–775. A review of the literatures indicates that only ba- Khatoon, S. and Ali, S. I. 1993. Chromosome Atlas of the Angio- sic chromosome numbers of x=6 and x=7 and mostly sperms of Pakistan. Department of Botany, University of Kara- 2n=14 have been counted in species of Scorzonera chi, Karachi. Krajevoy, S. U. 1934. On the question of chromosome variations in (Nazarova 1997, Safavi 1999, Martïn et al. 2012, Alti- Scorzonera tau-saghyz. Bot. J. 19: 367–375. (in Russian) nordu et al. 2015). Therefore the basic chromosome Martïn, E., Çetïn, Ö., Makbul, S., Duran, A., Öztürk, M., Boduroğlu, number in Scorzonera seems stable. We can conclude D. and Eşmekaya, B. 2012. Karyology of the Scorzonera L. (As- that x=10 for S. intricata is a rare and new basic chro- teraceae) taxa from Turkey. Turk. J. Biol. 36: 187–199. mosome number in Scorzonera. Mirzaghaderi, G. and Marzangi, K. 2015. IdeoKar: An ideogram constructing and karyotype analyzing software. Caryologia 68: Karyological parameters measured in S. intricata and 31–35. S. tortuosissima indicate almost a high degree of sym- 72 F. Bordbar et al. Cytologia 84(1)

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