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Method Article
Stefan L. Ameres
Institute of Molecular Biotechnology, IMBA, Vienna, Austria
Corresponding Author
Luisa Cochella
Research Institute of Molecular Pathology, IMP Vienna, Austria
Corresponding Author
License:
This work is licensed under a CC BY 4.0 License. Read Full License
The description of precise miRNA expression patterns is crucial to understand what these small RNAs contribute to animal development and physiology. High-throughput sequencing of miRNAs from individual developmental stages has provided insight into temporal regulation but often lacks the cellular resolution to link miRNA-function to the biology of distinct cell types within complex tissues. Here, we provide a protocol for microRNome by methylation-dependent sequencing (mime-seq), a chemical small RNA-tagging approach followed by chemoselective, high-throughput sequencing that enables the identification of tissue- and cell type-specific miRNA profiles in animals in a sensitive and reproducible manner. Using this strategy, we have uncovered the miRNA composition of specific cells and tissues within C. elegans and Drosophila, providing insights into miRNA spatial distribution at high resolution. This protocol accompanies Alberti et al (Nature Methods, published online February 26, 2017; 10.1038/nmeth.4610)
Keywords
miRNAs, small RNA methylation, HEN1, periodate oxidation, small RNA cloning, cell-specific miRNA profiling
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The authors declare no competing financial interests
This is a list of supplementary files associated with this preprint. Click to download.
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Table 7 List of primers, linkers and markers used in this protocol
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Associated Publications
Method Article
Stefan L. Ameres
Institute of Molecular Biotechnology, IMBA, Vienna, Austria
Corresponding Author
Luisa Cochella
Research Institute of Molecular Pathology, IMP Vienna, Austria
Corresponding Author
Version History
CURRENT STATUS: Posted
Version 1
Posted 12 Mar, 2018
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
The description of precise miRNA expression patterns is crucial to understand what these small RNAs contribute to animal development and physiology. High-throughput sequencing of miRNAs from individual developmental stages has provided insight into temporal regulation but often lacks the cellular resolution to link miRNA-function to the biology of distinct cell types within complex tissues. Here, we provide a protocol for microRNome by methylation-dependent sequencing (mime-seq), a chemical small RNA-tagging approach followed by chemoselective, high-throughput sequencing that enables the identification of tissue- and cell type-specific miRNA profiles in animals in a sensitive and reproducible manner. Using this strategy, we have uncovered the miRNA composition of specific cells and tissues within C. elegans and Drosophila, providing insights into miRNA spatial distribution at high resolution. This protocol accompanies Alberti et al (Nature Methods, published online February 26, 2017; 10.1038/nmeth.4610)
Figure 1
Figure 2
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Figure 9
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