BioSysBio:abstracts/2007/Stefan Weinges: Difference between revisions
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increased by more than 100% with respect to simple codon usage adaptation and several-fold with | increased by more than 100% with respect to simple codon usage adaptation and several-fold with | ||
respect to wild type sequence. | respect to wild type sequence. | ||
__NOTOC__ | __NOTOC__ |
Revision as of 19:36, 17 September 2006
=Computer Aided Multi-Parameter Gene Design: Impact of Synthetic DNAs on Protein Expression Enhancement=
Author(s): Frank Notka(1), Doris Leikam(1), Asli Bauer(2), David Raab(1), Marcus Graf(1) and Ralf Wagner (1,2)
Affiliations: (1)GENEART AG, Regensburg, (2) University of Regensburg, Molecular Microbiology and Gene Therapy
Contact:email: ralf.wagner@geneart.com
Keywords: 'add-keyword_1' 'add_keyword_2' 'add_keyword_3' 'add_keyword_4'
Protein production in cells is dependent on various factors including the underlying nucleotide sequence.
Gene optimization is dedicated to improve the expression properties of transgenes by codon adaptation to
the individual host, increasing RNA production, stability and nuclear export. However, most gene
optimization strategies depend on codon usage adaptation only, whereas RNA optimization relies on
optimization of many different parameters such as removal of RNA secondary structures, adjustment of
CG-values, avoidance of splice sites and elimination of instability elements. With the help of a multi
parameter optimizing algorithm, degeneration of the genetic code provides a powerful tool to identify,
analyze as well as utilize parameters and respective motifs to increase and/or adjust expression yields or
other important properties of a gene such as its safety or genetic stability.
Interestingly, in higher eukaryotes the overall CpG content was demonstrated to be crucial for the level of
transgene expression. In particular, we report here the intragenic CpG-dinucleotide dependent expression
pattern of differently designed synthetic genes, encoding human and murine cytokines, the green
fluorescence protein and HIV-1 proteins. Expression yields were monitored on the protein level for all
genetic variants and on the RNA level for GFP-recombinant cell lines. For all tested transiently transfected
and stable cell lines, a clear correlation of intragenic CpG-content, levels of cytoplasmic mRNA and
protein yields has been demonstrated. Using the maximum number of CpGs, expression yields could be
increased by more than 100% with respect to simple codon usage adaptation and several-fold with
respect to wild type sequence.