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Chromosomal detection of simple sequence repeats (SSRs) using nondenaturing FISH (ND-FISH)

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Abstract

Simple Sequence Repeats (SSRs) are known to be scattered and present in high number in eukaryotic genomes. We demonstrate that dye-labeled oligodeoxyribonucleotides with repeated mono-, di-, tri, or tetranucleotide motifs (15-20 nucleotides in length) have an unexpected ability to recognize SSR target sequences in non-denatured chromosomes. The results show that all these probes are able to invade chromosomes, independent of the size of the repeat motif, their nucleotide sequence, or their ability to form alternative B-DNA structures such as triplex DNA. This novel and remarkable property of binding SSR oligonucleotides to duplex DNA targets permitted the development of a non-denaturing fluorescence in situ hybridization method that quickly and efficiently detects SSR-enriched chromosome regions in mitotic, meiotic, and polytene chromosome spreads of different model organisms. These results have implications for genome analysis and for investigating the roles of SSRs in chromosome structure and function.

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Acknowledgments

This study was supported by grants from the Spanish Ministry of Science and Innovation (AGL 2009-10373). The authors thank Adrian Burton for linguistic assistance. We also wish to thank two anonymous referees for helpful comments and suggestions.

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Authors and Affiliations

  1. Department of Cell Biology and Genetics, University of Alcalá de Henares, 28871, Alcalá de Henares, Madrid, Spain

    Ángeles Cuadrado & Nicolás Jouve

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  1. Ángeles Cuadrado
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  2. Nicolás Jouve
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Correspondence to Ángeles Cuadrado.

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Communicated by L. Comai

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Figure S1

Confirmation that SSR-enriched chromosome regions in non-denatured barley chromosomes are mainly in a duplex state. The left and right panels show the results of ND-FISH (a-b) and FISH (c-d) experiments respectively when using (AAG)n as a probe. Only some interphase nuclei showed signals after ND-FISH (a-b) No signals were observed in mitotic chromosomes (b). Presumably, the main target structures detected using (AAG)n as probe, under nondenaturing hybridization conditions are ssDNA. In contrast, all the interphase nuclei (c) and mitotic chromosomes (d) showed signals after standard FISH. Scale bar 10 0 m (JPG 67.1 KB)

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Cuadrado, Á., Jouve, N. Chromosomal detection of simple sequence repeats (SSRs) using nondenaturing FISH (ND-FISH). Chromosoma 119, 495–503 (2010). https://doi.org/10.1007/s00412-010-0273-x

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