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Chromosomal study of a lamprey (Lampetra zanandreai Vladykov, 1955) (Petromyzonida: Petromyzontiformes): conventional and FISH analysis

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Abstract

Karyotype and other chromosomal characteristics in the Adriatic brook lamprey Lampetra zanandreai, representative of one of the most ancestral group of vertebrates, were examined using conventional (Ag-staining, C-banding as well as CMA3 and DAPI fluorescence) and molecular (FISH with 18/28S rDNA and EcoRI satDNA as probes) protocols with metaphase chromosomes derived from whole blood cultures. The chromosome complement had a modal diploid chromosome number of 2n = 164, as in other petromyzontid lamprey species. Ag-staining and CMA3 fluorescence, as well as FISH with 18/28S rDNA probes, detected nucleolar organizer regions (NORs) close to the centromeres of the biarmed chromosomes of pairs 1 and 2, the largest chromosome pairs of the complement. In addition to NORs, CMA3 fluorescence revealed positive signals in approximately 40 other chromosomes. DAPI stained mostly centromeric regions of many chromosomes as well as conspicuously massive blocks overlapping NOR sites. C-banding evidenced a large amount of constitutive heterochromatin in somatic chromosomes, with approximately 40 C-positive acrocentric elements completely heterochromatic, corresponding with the 40 CMA3+ chromosomes and positive heterochromatic blocks in pericentromeric regions of chromosome pairs 1 and 2. Polymerase chain reaction (PCR)-based cloning of satellite DNA with primers derived from Petromyzon marinus centromeric sequences was successful for L. zanandreai genomic DNA. The sequence was AT-rich (59%) and characterized by short consensus motifs similar to other centromeric satellite motifs. FISH using satDNA clones as a probe produced a fluorescent signal on a single pair of small chromosomes. This sequence was PCR-amplified also in L. planeri and P. marinus genomic DNA, and the evolution of this repetitive element in the above species was analysed.

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Abbreviations

CMA3 :

Chromomycin A3

DAPI:

4′,6-Diamidino-2 phenyl-indole

FISH:

Fluorescence in situ hybridization

K2P:

Kimura 2-parameters

NJ:

Neighbour-joining

NOR:

Nucleolar organizing region

rDNA:

Major ribosomal sites

satDNA:

Satellite DNA

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Acknowledgements

The authors are very grateful to Petr Ráb (Academy of the Sciences of the Czech Republic, Liběchov, Czech Republic) for valuable comments and suggestions on the manuscript. We wish to thank the provincial Administration of Macerata (Italy) for permitting specimen collection, Mario Marconi (University of Camerino, Italy) for assisting in field activities, and Mrs. Patricia O’Brien for linguistic revision. This research was financed by Università Politecnica delle Marche.

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Correspondence to Vincenzo Caputo.

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1

Southern blot hybridisation of the genomic DNA of L. zanandreai (lane 1), P. marinus (lane 2) and L. planeri (lane 3) digested with EcoRI. The hybridization was carried out with a digoxigenin-labelled probe from the centromeric satellite isolated from L. zanandreai by PCR. Arrow indicates the monomer of the EcoRI satDNA. (JPEG 85 kb)

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Caputo, V., Giovannotti, M., Cerioni, P.N. et al. Chromosomal study of a lamprey (Lampetra zanandreai Vladykov, 1955) (Petromyzonida: Petromyzontiformes): conventional and FISH analysis. Chromosome Res 19, 481–491 (2011). https://doi.org/10.1007/s10577-011-9197-4

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