Cross-site comparison of ribosomal depletion kits for Illumina RNAseq library construction
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Author(s)
Herbert, Zachary T
Kershner, Jamie P
Thimmapuram, Jyothi
Choudhari, Sulbha
Alekseyev, Yuriy O
Fan, Jun
Podnar, Jessica W
Wilcox, Edward
Gipson, Jenny
Gillaspy, Allison
Date Issued
March 2018
Journal
BMC Genomics
Publisher
BioMed Central Ltd
Citation
Herbert, Zachary T. et al. "Cross-site comparison of ribosomal depletion kits for Illumina RNAseq library construction" BMC Genomics 19, S2 (March 2018): 199 © 2018 The Author(s)
Version
Final published version
Abstract
Background
Ribosomal RNA (rRNA) comprises at least 90% of total RNA extracted from mammalian tissue or cell line samples. Informative transcriptional profiling using massively parallel sequencing technologies requires either enrichment of mature poly-adenylated transcripts or targeted depletion of the rRNA fraction. The latter method is of particular interest because it is compatible with degraded samples such as those extracted from FFPE and also captures transcripts that are not poly-adenylated such as some non-coding RNAs. Here we provide a cross-site study that evaluates the performance of ribosomal RNA removal kits from Illumina, Takara/Clontech, Kapa Biosystems, Lexogen, New England Biolabs and Qiagen on intact and degraded RNA samples.
Results
We find that all of the kits are capable of performing significant ribosomal depletion, though there are differences in their ease of use. All kits were able to remove ribosomal RNA to below 20% with intact RNA and identify ~ 14,000 protein coding genes from the Universal Human Reference RNA sample at >1FPKM. Analysis of differentially detected genes between kits suggests that transcript length may be a key factor in library production efficiency.
Conclusions
These results provide a roadmap for labs on the strengths of each of these methods and how best to utilize them. Keywords: RNAseqr; RNA depletion; Illumina; NGS; ABRF; Transcriptomics
MIT Department
Massachusetts Institute of Technology. Department of Biology
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Creative Commons Attribution
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DOI of Published Version
https://doi.org/10.1186/s12864-018-4585-1
