Molecular cytogenetic analysis of the crucian carp, Carassius carassius (Linnaeus, 1758) (Teleostei, Cyprinidae), using chromosome staining and fluorescence in situ hybridisation with rDNA probes.

Alicja Boron,Aneta Spoz,Lech Kirtiklis,Daisuke Ito,Syuiti Abe,Monika Karolewska,Dorota Juchno,Katarzyna Porycka

doi:10.3897/compcytogen.v8i3.7718

Abstract

The crucian carp Carassius carassius (Linnaeus, 1758) is a species with restricted and decreasing distribution in Europe. Six males and six females of the species from the Baltic Sea basin in Poland were examined to show sequentially CMA3/AgNO3 staining pattern, DAPI staining, and, for the first time in literature, molecular cytogenetic analysis using double-colour fluorescence in situ hybridisation (FISH) with 28S and 5S rDNA probes. The karyotype consisted of 20 m, 36 sm and 44 sta chromosomes, NF=156. The AgNO3 stained NORs were most frequently located terminally in the short arms of two sm and two sta elements, and CMA3-positive sites were also observed suggesting abundant GC-rich repetitive DNA in the regions. Other CMA3-positive sites in the short arms of six to ten sm and sta chromosomes were detected. The results based on 28S rDNA FISH confirmed the location of rDNA sites. DAPI-negative staining of NORs suggested the scarcity of AT-rich DNA in the regions. FISH with 5S rDNA probe revealed 8–14 loci (ten and 12 in respectively 49 and 29% of metaphases). They were located in two sm and eight to ten sta chromosomes and six of them were larger than others. Simultaneously, mapping of the two rDNA families on the chromosomes of C. carassius revealed that both 28S and 5S rDNA probes were located in different chromosomes. Molecular cytogenetic data of C. carassius presented here for the first time give an important insight into the structure of chromosomes of this polyploid and declining species and may be useful in its systematics.

Highlights

The genus Carassius Jarocki, 1882 is a fish group of polyploid origin as are some other cyprinids of subfamilies Cyprininae and Barbinae s.l., e.g. Cyprinus Linnaeus, 1758 and Barbus Cuvier, 1816 (Vasil’ev 1985, Le Comber and Smith 2004)
Chromosomes were counterstained with DAPI in Antifade solution (Vector Laboratories). We show here both single colour and dual colour fluorescence in situ hybridisation (FISH) with rDNAs because firstly prepared single colour FISH revealed DAPI banding pattern
After sequential staining with chromomycin A3 (CMA3), all signals were observed as a distinct bright fluorescence, suggesting abundant GC-rich repetitive DNA sequences in the regions (Fig. 1d)

Summary

Introduction

The genus Carassius Jarocki, 1882 is a fish group of polyploid origin as are some other cyprinids of subfamilies Cyprininae and Barbinae s.l., e.g. Cyprinus Linnaeus, 1758 and Barbus Cuvier, 1816 (Vasil’ev 1985, Le Comber and Smith 2004). The crucian carp, Carassius carassius (Linnaeus, 1758), native to Europe, is widely distributed from the northern France to the Danube drainage and Siberia, and from England in the north to the Alps in the south. This species is adapted to both a wide range of temperature and low oxygen content and prefers densely vegetated water bodies−backwaters and oxbows of lowland rivers, and lakes (Szczerbowski and Szczerbowski 2002, Freyhof and Kottelat 2008). In context of the genetic conservation of this species, it is important to determine its taxonomic diagnostic features possibly at all levels of its organisation including the chromosomal level

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