Karyological study on six Trigonella L. species (Leguminosae) in Turkey

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1 CARYOLOGIA Vol. 62, no. 2: , 2009 Karyological study on six Trigonella L. species (Leguminosae) in Turkey Yılmaz 1 Aykut, Esra Martin 2, Fatma Ünal 3 and Hasan Akan 4 1 Department of Biology, Faculty of Science and Arts, Abant İzzet Baysal University, Bolu, Turkey. 2 Department of Biology, Faculty of Education, Selçuk University, Konya, Turkey. 3 Department of Biology, Faculty of Science and Arts, Gazi University, Ankara, Turkey. 4 Department of Biology, Faculty of Science and Arts, Harran University, Şanlıurfa, Turkey. Abstract In this study, karyotype analyses of the six species of Trigonella L. were examined. Chromosome lengths were determined between µm in Trigonella astroites Fischer & C.Meyer, µm in Trigonella fischeriana Ser., µm in Trigonella velutina Boiss., µm in Trigonella cretica (L.) Boiss., µm in Trigonella coerulescens (M.Bieb.) Hal., µm in Trigonella procumbens (Bess.) Rchb., respectively. All species are diploid with 2n=14,16 and 18 chromosomes and the karyotypes of all species consist of metacentric chromosome pairs. Trigonella velutina has only satellites on the long arms of chromosome 2 and 6. The karyotype analysis of these species was first time counted in Turkey. Key words: karyotype, Trigonella, Turkey. INTRODUCTION The genus Trigonella is a member of Fabaceae family. Fabaceae is the second big family of flowering plants in the world with 650 genus and species (Ra k h e e et al. 2004). Trigonella L. is represented with about 100 species in the world and 50 species in Turkey (Se ç m e n et al. 1998). 50 taxa belonging to 13 sections are growing and localized in Mediterranean, Irano-Turanian and Europe-Siberian phytogeographical regions in Turkey. The total endemic taxa are 21, with 42% endemism rate (Hu b e r-mo r at h 1970; Dav i s et al. 1988; Ak a n et al. 2005; Ak a n et al. unpl. data 2006). The members of Trigonella are annual hairless or covered with feathers herbs and origins of them are the Mediterranean. Taxonomic classification of plant species is initially based on morphological characteristics. Because these features vary and sometimes difficult to observe, other taxonomic criteria such as chromosome number, chromosome morphology have been developed and these results have been used mostly in plant taxonomy. * Corresponding author: aykutyilmaza@gmail.com Here, we studied with six Trigonella species known as Trigonella cretica, Trigonella astroites, Trigonella fischeriana, Trigonella velutina, Trigonella coerulescens and Trigonella procumbens. The species analyzed in this study belonged to different Trigonella sections, such as Trigonella fischeriana and Trigonella astroites: section Bucerates, Trigonella coerulescens: section Biebersteinianae, Trigonella velutina: section Cylindricae, Trigonella cretica: section Samaroidea and Trigonella procumbens: section Capitatae (Hu b e r-mo r at h 1970). The present study was carried out to determine chromosome numbers, types, arm lengths according to centromer position and other detailed measurements of six Trigonella species. Among the studied species, Trigonella astroites, Trigonella coerulescens, Trigonella velutina and Trigonella cretica have diploid chromosome with 2n=16, Trigonella procumbens has diploid chromosome with 2n=18 and finally Trigonella fischeriana has diploid chromosome with 2n=14. MATERIALS AND METHODS The materials used in this study were collected in different areas of Turkey. The localities, collectors and species names are shown in Table 1.

2 90 yilmaz, martin, ünal and akan Ta b l e 1 Localities of species used in the study. Species Localities Collector name and number Trigonella astroites Between Kayseri-Ürgüp, edges of road, 29.km.,1180m. Akan 3771 Trigonella fischeriana Kayseri: Ali mountain, North-east edges, 1350m. Akan 3654 Trigonella velutina Hadim-Konya, edges of road, 10.km.,1350m. Akan 3790 Trigonella cretica Burdur: Gölhisar-Dirmil, 5.km., m. Akan 3480 Trigonella coerulescens Kayseri:Hisarcık-Erciyes mountain road, near to Hisarcık, 1550m. Akan 3659 Trigonella procumbens Akşehir-Gelendost,1.5km.,edges of road, 1110km. Akan 4760 For karyotype studies, firstly; seeds of collected species were put on moist filter paper in petri dishes at room temperature. Germinated seeds having root tips in lengths of 2-10 mm. are taken and put into tubes. As a first treatment, root tips were transferred into tubes and were pretreated in α-monobromonaphtalene for hours at 4 o C. Then, roots were fixed in 3:1 absolute alcohol-acetic acid for over night. Hydrolization process was done in 1N HCL at 60 o C for changeable times according to species. Root tips were stained with giemsa for 2 hours. Squashes were made in % 45 acetic acid. After removing of cover slips in liquid nitrogen, slides were dried in air and then mounted in Depex. Photographs were taken using LEICA DM LB2 microscope with camera. The relative lengths of species were calculated according to the nomenclature of El ç i (1982) and chromosomes were classified according to the nomenclature of Leva n et al. (1964). Fig. 1 Somatic chromosomes in Trigonella. a) Trigonella astroites, b) Trigonella coerulescens, c) Trigonella cretica, d) Trigonella velutina, e) Trigonella fischeriana, f) Trigonella procumbens. Bar: 10 µm.

3 karyological study on six trigonella l. species (leguminosae) in turkey 91 Ta b l e 2 Measurements (μm) of somatic chromosomes of five Trigonella species. Chromosome no Chromosome arms Long arm (L) (μm) Short arm (S) (μm) Satellite (μm) Arm ratio (L/S) Total Length (μm) Relative Length (%) Chromosome type T. astroites m m m m m m m m Haploid chromosome lenght: μm T. coerulescens m m m m m m m m Haploid chromosome lenght: μm T. cretica m m m m m m m m Haploit chromosome lenght: 9.96 μm T. velutina m m m m m m m m Haploit chromosome lenght: μm T. fischeriana m m m m m m m Haploit chromosome lenght: 8.46 μm

4 92 yilmaz, martin, ünal and akan RESULTS Karyotype analyses of the six species of Trigonella were determined but detailed karyotype analyses were only made on five species of Trigonella (Trigonella astroites, Trigonella coerulescens, Trigonella cretica, Trigonella velutina and Trigonella fischeriana). Trigonella procumbens was only observed according to chromosome number, total lengths of chromosomes and haploid chromosome length. All somatic cells of investigated species have diploid chromosomes with 2n=14, 16 and 18. While chromosome numbers of Trigonella astroites, Trigonella coerulescens, Trigonella cretica, Trigonella velutina are 2n=16, Trigonella fischeriana and Trigonella procumbens have 2n=14 and 2n=18 chromosomes, respectively. Averages of chromosomal lengths of investigated species ranged from 0.98 to 2.59 μm. Among these, while Trigonella cretica has the smallest chromosome (Fig. 1-c), the biggest chromosome was observed on Trigonella velutina (Fig. 1-d). Detailed measurements of somatic chromosomes of five Trigonella species are given in Table 2. Among these, Trigonella fischeriana has the smallest haploid chromosome length. However, Trigonella fischeriana consist of n=7 haploid chromosomes, as other four species have n=8 haploid chromosomes. Haploid complement values are very close in Trigonella astroites, Trigonella coerulescens and Trigonella cretica. Trigonella velutina has bigger haploid complement than other species. The differences may be derived from satellites located on Trigonella velutina. Fundamentally, most of the studied species showed very similar karyotypic characters. Distinctions observed may be caused from different chromosome numbers. DISCUSSION In this study, we determined chromosome numbers and detailed measurements of six species of Trigonella as 2n=14,16 and 18 for the first time in Turkey. When the results provided from this study are compared with previous studies on the chromosome numbers of Trigonella taxa, they do not show much variation except Trigonella procumbens. We determined chromosome number of Trigonella procumbens as 2n=18. There are not enough literature data on chromosome numbers and karyotype of Trigonella species. Previous reports show that all the investigated members of Trigonella have diploid chromosome with 2n=14 and 16 (Bi d a k and Am i n 1996; Da s et al. 2000; Ia n et al. 2006). This study together with former results show that basic haploid chromosome numbers of Trigonella may be n=7,8 and 9. Although species investigated for karyotype analysis in this study have small chromosomes, karyotype analyses of the six species of Trigonella were examined. Differences in chromosome numbers and total lengths might result with an addition or deletion of DNA (Na r aya n and Du r r a n t 1983). In addition, species that were studied here, have been found within five different sections and for this reason chromosome numbers and lengths can be different. Comprehensive study involving different section and species of Trigonella in Turkey is necessary to understand Trigonella taxonomy effectively. This study on the karyotype of six Trigonella species showed that all chromosomes are metacentric and among these species, Trigonella velutina has only satellite. Four of the studied species have the same chromosome number with 2n=16. Other species have 2n=14 and 2n=18 chromosome. At the same time, among the studied species Trigonella velutina show little distinction Ta b l e 3 Measurements (μm) of somatic chromosomes of Trigonella procumbens. Chromosome no Total length (μm) Relative length (%) Haploid chromosome length: μm

5 karyological study on six trigonella l. species (leguminosae) in turkey 93 Fig. 2 Idiograms of five species of Trigonella. a) Trigonella astroites, b) Trigonella coerulescens, c) Trigonella cretica, d) Trigonella velutina, e) Trigonella fischeriana.

6 94 yilmaz, martin, ünal and akan according to chromosome length. Chromosome lengths of other species are very similar and this makes Trigonella velutina slightly different from the other species. Acknowledgements The author thanks to Biology Department of Abant İzzet Baysal University for providing support. REFERENCES Ak a n H., Ek i c i M. and Ay ta ç Z., 2005 The endemic species of Trigonella and their conservation in Turkey. XVII. International Botanical Congress, Vienna: (17-23 July), Abstracts, p Ak a n H., Ek ı c ı M., Ay ta ç Z. and Pı n a r M.N., 2006 The Taxonomic Revision of Trigonella L. In Turkey. The Project of The Scientific and Technological Research Council of Turkey (TÜBİTAK, Project No: TBAG-2099), Ankara: Unpublished data. Bı d a k A.B. and Am ı n A.W., 1996 Inter and Intraspesific Chromosomal Variations in Four Species of Trigonella. J. Union Arab Biol., 3(B): Da s A.B., Mo h a n t y S., Th a n g a r a j T. and Da s P., 2000 Variation of 4C DNA Content and Karyotype in Nine Cultivars of Fenugreek (Trigonella foenumgraecum L.). Journal of Herbs, Spices & Medicinal Plants, 7:1,25-32 Dav ı s P.H., Mı l l R.R. and Ta n K. (eds.)., 1988 Flora of Turkey and the East Aegean Islands (supplement 1). Edinburgh: Edinburgh University Press. El ç ı Ş., 1982 Sitogenetikte Gözlemler ve Araştırma Yöntemleri. Uğurel matbaası-malatya, pp Hu b e r-mo r at h A., 1970 Trigonella L. In Davis PH (ed.), Flora of Turkey and The East Aegean Islands. 3: Edinburgh: Edinburgh Univ. Press. Ia n S.D., Ra m a k r ı s h n a n M.N. and Daw n C.V., 2006 First report of meiotic chromosome number and karyotype analysis of an accession of T. balansae (Leguminosae). New Zealand Journal of Agricultural Research, 49: Le va n A., Fr e d g a K. and Sa n d b e r g A.A., 1964 Nomenclature for centromeric position on chromosomes. Hereditas, 52: Na r aya n R.K.J. and Du r r a n t A., 1983 DNA distribution in chromosomes of Lathyrus species. Genetica, 61: Ra k h e e S.D., Me e n a D.L., La l B.C., Pr a b h a k a r K.R. and Vı d ya S.G., 2004 Assesment of genetic diversity in Trigonella foenum-graecum and Trigonella caerulea using ISSR and RAPD markers. BMC Plant Biology, 4/13. Se ç m e n Ö.,Ge m ı c ı Y., Gö r k G., Be k at L. and Le b l e b ı c ı E., 1998 Tohumlu Bitkiler Sistematiği. Bornova- İzmir, Received May 21 th 2009; accepted February 15 th 2009