Abstract
Repetitive DNAs comprise large portion of eukaryote genomes. In genome projects, the assembly of repetitive DNAs is challenging due to the similarity between repeats, which generate ambiguities for alignment. Fluorescence in situ hybridization (FISH) is a powerful technique for the physical mapping of various sequences on chromosomes. This technique is thus very helpful in chromosome-based genome assemblies, providing information on the fine architecture of genomes and their evolution. However, various protocols are currently used for FISH mapping, most of which are relatively laborious and expensive, or work properly only with a specific type of probes or sequences, and there is a need for a universal and affordable FISH protocol. Here we tested a FISH protocol for mapping of different DNA repeats, such as multigene families (rDNAs, U snDNAs, histone genes), satellite DNAs, microsatellites, transposable elements, DOP-PCR products, and telomeric motif (TTAGG)n, on the chromosomes of various insects and other arthropods. Different cell types and stages obtained from diverse tissues were used. The FISH procedure proved high quality and reliable results in all experiments performed. We obtained data on the chromosomal distribution of DNA repeats in representatives of insects and other arthropods. Thus, our results allow us to conclude that the protocol is universal and requires only time adjustment for chromosome/DNA denaturation. The use of this FISH protocol will facilitate studies focused on understanding the evolution and role of repetitive DNA in arthropod genomes.
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Acknowledgements
We are grateful to a number of colleagues for providing samples of insects, other arthropods, and nematodes (Allison K. Anjos and Vanessa B. Bardella, Rio Claro, SP, Brazil; Rita C. Moura, Recife, PE, Brazil; Jack E. Green, Cambridge, UK; David G. Heckel and Melanie Unbehend, Jena, Germany; Jiří Král, Prague, Czech Republic; Daniela Hlávková, Dalibor Kodrík, Marie Korchová, Petr Nguyen, Vladimír Půža, Pavlína Větchová, and Magda Zrzavá, all České Budějovice, Czech Republic), determination of the sawfly Nematus ribesii (Libor Dvořák, Mariánské Lázně, Czech Republic), providing some images (Camila C. Avelino, Mara M.L.S. Pinheiro, and Octavio M. Palacios-Gimenez, all São Paulo, SP, Brazil), and performing some experiments (Vanessa B. Bardella, Mara M.L.S. Pinheiro, Octavio M. Palacios-Gimenez). We also wish to thank members of FM laboratory (Atsuo Yoshido, Magda Zrzavá, Anna Voleníková, Martina Hejníčková, and Sander Visser) for valuable suggestions on the FISH protocol. Last but not least, we thank Tatiane C. Mariguela (Rio Claro, SP, Brazil) for schematic drawings of insects in Figure S1. This study was supported by CNPq-Conselho Nacional de Pesquisa e Desenvolvimento, FAPESP-Fundação de Amparo à Pesquisa do Estado de São Paulo (BE.PQ, process number 2017/097319, Regular Research Grant, process number 2019/19069-7) and by grants 17-13713S and 20-13784S of the Czech Science Foundation given to FM.
Funding
CNPq-Conselho Nacional de Pesquisa e Desenvolvimento, FAPESP-Fundação de Amparo à Pesquisa do Estado de São Paulo (BE.PQ, process number 2017/097319, Regular Research Grant, process number 2019/19069-7 given to DCCM) and by grants 17-13713S and 20-13784S the Czech Science Foundation given to FM.
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Cabral-de-Mello, D.C., Marec, F. Universal fluorescence in situ hybridization (FISH) protocol for mapping repetitive DNAs in insects and other arthropods. Mol Genet Genomics 296, 513–526 (2021). https://doi.org/10.1007/s00438-021-01765-2
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DOI: https://doi.org/10.1007/s00438-021-01765-2