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Efficient cellular ...
Efficient cellular fractionation improves RNA sequencing analysis of mature and nascent transcripts from human tissues
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- Zaghlool, Ammar, 1980- (author)
- Uppsala universitet,Institutionen för immunologi, genetik och patologi,Science for Life Laboratory, SciLifeLab,Lars Feuk's group
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- Ameur, Adam (author)
- Uppsala universitet,Institutionen för immunologi, genetik och patologi,Science for Life Laboratory, SciLifeLab
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- Nyberg, Linnea (author)
- Uppsala universitet,Institutionen för immunologi, genetik och patologi,Science for Life Laboratory, SciLifeLab
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- Halvardson, Jonatan (author)
- Uppsala universitet,Institutionen för immunologi, genetik och patologi,Science for Life Laboratory, SciLifeLab
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- Grabherr, Manfred (author)
- Uppsala universitet,Institutionen för medicinsk biokemi och mikrobiologi,Science for Life Laboratory, SciLifeLab
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- Cavelier, Lucia (author)
- Uppsala universitet,Institutionen för immunologi, genetik och patologi,Science for Life Laboratory, SciLifeLab
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- Feuk, Lars (author)
- Uppsala universitet,Institutionen för immunologi, genetik och patologi,Science for Life Laboratory, SciLifeLab
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(creator_code:org_t)
- 2013-11-13
- 2013
- English.
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In: BMC Biotechnology. - : Springer Science and Business Media LLC. - 1472-6750. ; 13, s. 99-
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https://doi.org/10.1...
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https://uu.diva-port... (primary) (Raw object)
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https://bmcbiotechno...
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https://urn.kb.se/re...
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Abstract
Subject headings
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- Background: The starting material for RNA sequencing (RNA-seq) studies is usually total RNA or polyA+ RNA. Both forms of RNA represent heterogeneous pools of RNA molecules at different levels of maturation and processing. Such heterogeneity, in addition to the biases associated with polyA+ purification steps, may influence the analysis, sensitivity and the interpretation of RNA-seq data. We hypothesize that subcellular fractions of RNA may provide a more accurate picture of gene expression. Results: We present results for sequencing of cytoplasmic and nuclear RNA after cellular fractionation of tissue samples. In comparison with conventional polyA+ RNA, the cytoplasmic RNA contains a significantly higher fraction of exonic sequence, providing increased sensitivity in expression analysis and splice junction detection, and in improved de novo assembly of RNA-seq data. Conversely, the nuclear fraction shows an enrichment of unprocessed RNA compared with total RNA-seq, making it suitable for analysis of nascent transcripts and RNA processing dynamics. Conclusion: Our results show that cellular fractionation is a more rapid and cost effective approach than conventional polyA+ enrichment when studying mature RNAs. Thus, RNA-seq of separated cytosolic and nuclear RNA can significantly improve the analysis of complex transcriptomes from mammalian tissues.
Subject headings
- NATURVETENSKAP -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)
- NATURAL SCIENCES -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)
Keyword
- Cuyoplasmic RNA
- Nuclear RNA
- RNA sequencing
- mRNA
Publication and Content Type
- ref (subject category)
- art (subject category)
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