Characterization of c-banded mitotic chromosomes in four species of anuran amphibians from Thailand

Abstract

Short Notes Characterization of c-banded mitotic chromosomes in four species of anuran amphibians from Thailand Thavorn Supaprom 1 , Visut Baimai 2 Chromosomes of anuran amphibians are par- ticularly interesting in terms of their markedly conservative chromosome number and mor- phology. However, they generally show extra- ordinary variation in the distribution and mor- phology of secondary constrictions. The karyol- ogy of anuran amphibians has been investigated predominantly by conventionally stained chro- mosomes and a careful comparative karyotypic analysis has been made among individuals of some anuran families (Seto, 1965; Morescalchi, 1973). Criteria used for comparison include arm ratio, relative length and centromeric in- dex as well as chromosome number and posi- tion of the secondary constriction (Blommer- Schlosser, 1978; Supaprom and Baimai, 2002, 2003). In order to characterise anuran chro- mosomes in more detail, differential staining techniques have been found useful for multiple bands as well as for indicating the molecular structure of the chromosomes (Schmid, 1978b). Some authors have already reported heterochro- matin banding patterns in chromosomes of cer- tain groups of anuran amphibians (Stock and Mengden, 1975; Schmid, 1978a, 1978b; King, 1980; Birstein and Mazin, 1982; Matsui et al., 1 - Department of Biological Science, Faculty of Science, Ubon Ratchathani University, Warin-Dejudom Road, Warinchamrap, Ubon Ratchathani 34190, Thailand 2 - Department of Biology, Faculty of Science, Mahidol University, Rama VI Road, Bangkok 10400, Thailand Corresponding author; e-mail: scvbm@mahidol.ac.th 1985). The c-bands are known to contain re- peated sequences of DNA and can be classified into three groups: centromeric, interstitial and telomeric regions (King, 1991). In this paper, the distribution of constitu- tive heterochromatin in the form of positive c- banding occurring in the somatic chromosomes of 4 species of anuran amphibians from Thai- land are described. Materials and methods Taxonomy and chromosome preparation. The four species of frog were Rana lateralis Boulenger 1887, Limnonectes pileatus (Boulenger, 1916) Glyphoglossus molossus Gun- ther 1868, and Polypedates leucomystax (Gravenhorst, 1829). Twenty individuals of these species of frog were collected from a variety of natural habitats in Northeast- ern and Northern Thailand (See Appendix). The specimens were brought back to the laboratory at Ubon Ratchathani University for chromosomal analysis. The specimens were identified using the methods of Taylor (1962) and Chan-Ard et al. (1999). Preserved specimens are deposited at the De- partment of Biological Science, Faculty of Science, Ubon Ratchathani University. Mitotic chromosomes were prepared directly from bone marrow cells after in vivo colcemid treatment (0.3%). The preparation of hypotonic cell suspensions and chromosome fixation followed the technique of Schmid (1978a) with slight modifications as follows. The bone marrow was taken from limb bones. After the cartilageneous epiphyses were cut off from the limb bones, the bone marrow was flushed out using a fine hypodermic needle containing 10 ml of hy- potonic KCl solution (0.075 M) into a centrifuge tube. The bone marrow was then vigorously resuspended using a Pas- teur pipette and left in the hypotonic KCl solution at 37 ◦ C for 20 min. After hypotonic treatment, the cell suspension © Koninklijke Brill NV, Leiden, 2005. Amphibia-Reptilia 26 (2005): 367-371 Also available online - www.brill.nl