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Parallel DNA pyrosequencing unveils new zebrafish microRNAs
BACKGROUND: MicroRNAs (miRNAs) are a new class of small RNAs of approximately 22 nucleotides in length that control eukaryotic gene expression by fine tuning mRNA translation. They regulate a wide variety of biological processes, namely developmental timing, cell differentiation, cell proliferation,...
Autores principales: | , , , , , , , , |
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Formato: | Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2009
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2684549/ https://www.ncbi.nlm.nih.gov/pubmed/19397817 http://dx.doi.org/10.1186/1471-2164-10-195 |
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author | Soares, Ana R Pereira, Patrícia M Santos, Bruno Egas, Conceição Gomes, Ana C Arrais, Joel Oliveira, José L Moura, Gabriela R Santos, Manuel AS |
author_facet | Soares, Ana R Pereira, Patrícia M Santos, Bruno Egas, Conceição Gomes, Ana C Arrais, Joel Oliveira, José L Moura, Gabriela R Santos, Manuel AS |
author_sort | Soares, Ana R |
collection | PubMed |
description | BACKGROUND: MicroRNAs (miRNAs) are a new class of small RNAs of approximately 22 nucleotides in length that control eukaryotic gene expression by fine tuning mRNA translation. They regulate a wide variety of biological processes, namely developmental timing, cell differentiation, cell proliferation, immune response and infection. For this reason, their identification is essential to understand eukaryotic biology. Their small size, low abundance and high instability complicated early identification, however cloning/Sanger sequencing and new generation genome sequencing approaches overcame most technical hurdles and are being used for rapid miRNA identification in many eukaryotes. RESULTS: We have applied 454 DNA pyrosequencing technology to miRNA discovery in zebrafish (Danio rerio). For this, a series of cDNA libraries were prepared from miRNAs isolated at different embryonic time points and from fully developed organs. Each cDNA library was tagged with specific sequences and was sequenced using the Roche FLX genome sequencer. This approach retrieved 90% of the 192 miRNAs previously identified by cloning/Sanger sequencing and bioinformatics. Twenty five novel miRNAs were predicted, 107 miRNA star sequences and also 41 candidate miRNA targets were identified. A miRNA expression profile built on the basis of pyrosequencing read numbers showed high expression of most miRNAs throughout zebrafish development and identified tissue specific miRNAs. CONCLUSION: This study increases the number of zebrafish miRNAs from 192 to 217 and demonstrates that a single DNA mini-chip pyrosequencing run is effective in miRNA identification in zebrafish. This methodology also produced sufficient information to elucidate miRNA expression patterns during development and in differentiated organs. Moreover, some zebrafish miRNA star sequences were more abundant than their corresponding miRNAs, suggesting a functional role for the former in gene expression control in this vertebrate model organism. |
format | Text |
id | pubmed-2684549 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2009 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-26845492009-05-20 Parallel DNA pyrosequencing unveils new zebrafish microRNAs Soares, Ana R Pereira, Patrícia M Santos, Bruno Egas, Conceição Gomes, Ana C Arrais, Joel Oliveira, José L Moura, Gabriela R Santos, Manuel AS BMC Genomics Research Article BACKGROUND: MicroRNAs (miRNAs) are a new class of small RNAs of approximately 22 nucleotides in length that control eukaryotic gene expression by fine tuning mRNA translation. They regulate a wide variety of biological processes, namely developmental timing, cell differentiation, cell proliferation, immune response and infection. For this reason, their identification is essential to understand eukaryotic biology. Their small size, low abundance and high instability complicated early identification, however cloning/Sanger sequencing and new generation genome sequencing approaches overcame most technical hurdles and are being used for rapid miRNA identification in many eukaryotes. RESULTS: We have applied 454 DNA pyrosequencing technology to miRNA discovery in zebrafish (Danio rerio). For this, a series of cDNA libraries were prepared from miRNAs isolated at different embryonic time points and from fully developed organs. Each cDNA library was tagged with specific sequences and was sequenced using the Roche FLX genome sequencer. This approach retrieved 90% of the 192 miRNAs previously identified by cloning/Sanger sequencing and bioinformatics. Twenty five novel miRNAs were predicted, 107 miRNA star sequences and also 41 candidate miRNA targets were identified. A miRNA expression profile built on the basis of pyrosequencing read numbers showed high expression of most miRNAs throughout zebrafish development and identified tissue specific miRNAs. CONCLUSION: This study increases the number of zebrafish miRNAs from 192 to 217 and demonstrates that a single DNA mini-chip pyrosequencing run is effective in miRNA identification in zebrafish. This methodology also produced sufficient information to elucidate miRNA expression patterns during development and in differentiated organs. Moreover, some zebrafish miRNA star sequences were more abundant than their corresponding miRNAs, suggesting a functional role for the former in gene expression control in this vertebrate model organism. BioMed Central 2009-04-27 /pmc/articles/PMC2684549/ /pubmed/19397817 http://dx.doi.org/10.1186/1471-2164-10-195 Text en Copyright © 2009 Soares et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( (http://creativecommons.org/licenses/by/2.0) ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Article Soares, Ana R Pereira, Patrícia M Santos, Bruno Egas, Conceição Gomes, Ana C Arrais, Joel Oliveira, José L Moura, Gabriela R Santos, Manuel AS Parallel DNA pyrosequencing unveils new zebrafish microRNAs |
title | Parallel DNA pyrosequencing unveils new zebrafish microRNAs |
title_full | Parallel DNA pyrosequencing unveils new zebrafish microRNAs |
title_fullStr | Parallel DNA pyrosequencing unveils new zebrafish microRNAs |
title_full_unstemmed | Parallel DNA pyrosequencing unveils new zebrafish microRNAs |
title_short | Parallel DNA pyrosequencing unveils new zebrafish microRNAs |
title_sort | parallel dna pyrosequencing unveils new zebrafish micrornas |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2684549/ https://www.ncbi.nlm.nih.gov/pubmed/19397817 http://dx.doi.org/10.1186/1471-2164-10-195 |
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