Format

Send to

Choose Destination
PLoS One. 2011 Mar 3;6(3):e17596. doi: 10.1371/journal.pone.0017596.

Multiparameter RNA and codon optimization: a standardized tool to assess and enhance autologous mammalian gene expression.

Author information

1
Geneart/Life Technologies, [corrected] BioPark, Regensburg, Germany.

Erratum in

  • PLoS One. 2011;6(3). doi: 10.1371/annotation/039deb02-bbe7-406c-a876-341cc4f3fefa.

Abstract

Autologous expression of recombinant human proteins in human cells for biomedical research and product development is often hampered by low expression yields limiting subsequent structural and functional analyses. Following RNA and codon optimization, 50 candidate genes representing five classes of human proteins--transcription factors, ribosomal and polymerase subunits, protein kinases, membrane proteins and immunomodulators--all showed reliable, and 86% even elevated expression. Analysis of three representative examples showed no detrimental effect on protein solubility while unaltered functionality was demonstrated for JNK1, JNK3 and CDC2 using optimized constructs. Molecular analysis of a sequence-optimized transgene revealed positive effects at transcriptional, translational, and mRNA stability levels. Since improved expression was consistent in HEK293T, CHO and insect cells, it was not restricted to distinct mammalian cell systems. Additionally, optimized genes represent powerful tools in functional genomics, as demonstrated by the successful rescue of an siRNA-mediated knockdown using a sequence-optimized counterpart. This is the first large-scale study addressing the influence of multiparameter optimization on autologous human protein expression.

PMID:
21408612
PMCID:
PMC3048298
DOI:
10.1371/journal.pone.0017596
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Public Library of Science Icon for PubMed Central
Loading ...
Support Center