Targeted enrichment outperforms other enrichment techniques and enables more multi-species RNA-Seq analyses

• Published in: Scientific reports 2018-09, Vol.8 (1), p.13377-12, Article 13377
• Main Authors: Chung, Matthew, Teigen, Laura, Liu, Hong, Libro, Silvia, Shetty, Amol, Kumar, Nikhil, Zhao, Xuechu, Bromley, Robin E., Tallon, Luke J., Sadzewicz, Lisa, Fraser, Claire M., Rasko, David A., Filler, Scott G., Foster, Jeremy M., Michalski, Michelle L., Bruno, Vincent M., Dunning Hotopp, Julie C.
• Format: Article
• Language: English
• Subjects: 38/39; 45/91; 631/208/212/2019; 631/326/417/2551; 631/326/421; Enrichment; Gene mapping; Humanities and Social Sciences; Infections; Medical research; Medicine; Mosquitoes; multidisciplinary; Organisms; Pathogens; Peptide mapping; Polyadenine; Science; Science (multidisciplinary); Species; Tropical diseases
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-018-31420-7

Enrichment methodologies enable the analysis of minor members in multi-species transcriptomic data. We compared the standard enrichment of bacterial and eukaryotic mRNA to a targeted enrichment using an Agilent SureSelect (AgSS) capture for Brugia malayi , Aspergillus fumigatus , and the Wolbachia endosymbiont of B. malayi ( w Bm). Without introducing significant systematic bias, the AgSS quantitatively enriched samples, resulting in more reads mapping to the target organism. The AgSS-enriched libraries consistently had a positive linear correlation with their unenriched counterparts (r 2  = 0.559–0.867). Up to a 2,242-fold enrichment of RNA from the target organism was obtained following a power law (r 2  = 0.90), with the greatest fold enrichment achieved in samples with the largest ratio difference between the major and minor members. While using a single total library for prokaryote and eukaryote enrichment from a single RNA sample could be beneficial for samples where RNA is limiting, we observed a decrease in reads mapping to protein coding genes and an increase in multi-mapping reads to rRNAs in AgSS enrichments from eukaryotic total RNA libraries compared to eukaryotic poly(A)-enriched libraries. Our results support a recommendation of using AgSS targeted enrichment on poly(A)-enriched libraries for eukaryotic captures, and total RNA libraries for prokaryotic captures, to increase the robustness of multi-species transcriptomic studies.