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Sep 14, 2009

DNA2.0 Reveals Intricacies Involved in Optimized Gene Design

  • DNA2.0 reports that is has figured out how to chose codons that will optimize protein expression. The company says that the set of design algorithms it has developed enables the firm to offer expression yields up to 50 times greater than competing approaches.

    The research is published in PLoS ONE in a paper titled “Design Parameters to Control Synthetic Gene Expression in Escherichia coli.” The results are contrary to some of the most widely held assumptions, according to the firm. In particular, it found that the common practice of using the codons that are used most highly in the native genes of an organism can actually reduce expression.

    To achieve high levels of protein expression, many researchers have abandoned cloning in favor of synthetic genes, which can be acquired in a couple of weeks at a reasonable cost. Unlike a natural gene, a synthetic gene can be designed to use any codon to encode each amino acid. This has the potential to be a huge advantage, as most researchers have come to accept that codon optimization can affect protein expression.

    Since a protein can be encoded by many alternative nucleic acid sequences, however, a gene design strategy is required to predict the sequence that will result in optimal expression. Several different approaches to optimization, including codon sampling, codon pair optimization, and codon frequency matching, have been based primarily on analysis of genomic sequences. These have proven to be hit-or-miss, according to DNA2.0.

    The company’s current research has uncovered the elements in gene design that determine the protein-expression yields that can be obtained from a DNA sequence. “Our systematic analysis of gene-design parameters in this study has allowed us to identify codon usage within a gene as a critical determinant of protein-expression levels in E. coli,” says Mark Welch, Ph.D., director of gene design for DNA2.0 and author of the paper. “We propose a biochemical basis for this as well as design algorithms to ensure high protein production from synthetic genes. Replication of this methodology has already allowed us to derive design algorithms for several additional expression systems.”

    Researchers at DNA2.0 synthesized two sets of about 40 genes, one encoding phi29 polymerase and the other a single-chain antibody.  Expression of the genes was measured in E. coli. Levels ranged from undetectable to 30% of cellular protein. Correlation of sequences with expression levels led to the conclusion that protein expression strongly depends on codon choice.

    Surprisingly, the favored codons were not those most abundant in highly expressed native E. coli genes. Rather they were those codons read by tRNAs that are most highly charged during amino acid starvation.

    “DNA2.0 has discovered the Rosetta stone for determining the best codon choices, which means that we can design genes for our customers that are guaranteed for high-expression yields,” claims Jeremy Minshull, president of DNA2.0. “The PLoS ONE publication nicely complements the broad and fundamental patents we were recently awarded for this technology; peer review is essential for scientific credibility, and none of our competitors can come close to this."

     



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