Next-generation sequencing of the Chinese hamster ovary microRNA transcriptome: Identification, annotation and profiling of microRNAs as targets for cellular engineering

Matthias Hackl, Tobias Jakobi, Jochen Blom, Daniel Doppmeier, Karina Brinkrolf, Rafael Szczepanowski, Stephan H. Bernhart, Christian Höner zu Siederdissen, Juan A.Hernandez Bort, Matthias Wieser, Renate Kunert, Simon Jeffs, Ivo L. Hofacker, Alexander Goesmann, Alfred Pühler, Nicole Borth, Johannes Grillari

Research output: Contribution to journalArticlepeer-review

83 Scopus citations

Abstract

Chinese hamster ovary (CHO) cells are the predominant cell factory for the production of recombinant therapeutic proteins. Nevertheless, the lack in publicly available sequence information is severely limiting advances in CHO cell biology, including the exploration of microRNAs (miRNA) as tools for CHO cell characterization and engineering. In an effort to identify and annotate both conserved and novel CHO miRNAs in the absence of a Chinese hamster genome, we deep-sequenced small RNA fractions of 6 biotechnologically relevant cell lines and mapped the resulting reads to an artificial reference sequence consisting of all known miRNA hairpins. Read alignment patterns and read count ratios of 5' and 3' mature miRNAs were obtained and used for an independent classification into miR/miR* and 5p/3p miRNA pairs and discrimination of miRNAs from other non-coding RNAs, resulting in the annotation of 387 mature CHO miRNAs. The quantitative content of next-generation sequencing data was analyzed and confirmed using qPCR, to find that miRNAs are markers of cell status. Finally, cDNA sequencing of 26 validated targets of miR-17-92 suggests conserved functions for miRNAs in CHO cells, which together with the now publicly available sequence information sets the stage for developing novel RNAi tools for CHO cell engineering.

Original languageEnglish (US)
Pages (from-to)62-75
Number of pages14
JournalJournal of Biotechnology
Volume153
Issue number1-2
DOIs
StatePublished - Apr 20 2011
Externally publishedYes

Keywords

  • Chinese hamster ovary cells
  • MicroRNA
  • Next-generation sequencing

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

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