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Biomedical subjects

Stephen J Freeland

Publications and source records attributed to Stephen J Freeland.

6 recordsLinked to original sources
PubMed2006

SGDB: a database of synthetic genes re-designed for optimizing protein over-expression.

Here we present the Synthetic Gene Database (SGDB): a relational database that houses sequences and associated experimental information on synthetic (artificially engineered) genes from all peer-reviewed studies published to date. At present, the database comprises information from more than 200 published experiments. This resource not only provides reference material to guide experimentalists in designing new genes that improve protein expression, but also offers a dataset for analysis by bioinformaticians who seek to test ideas regarding the underlying factors that influence gene expression. The SGDB was built under MySQL database management system. We also offer an XML schema for standardized data description of synthetic genes. Users can access the database at http://www.evolvingcode.net/codon/sgdb/index.php, or batch downloads all information through XML files. Moreover, users may visually compare the coding sequences of a synthetic gene and its natural counterpart with an integrated web tool at http://www.evolvingcode.net/codon/sgdb/aligner.php, and discuss questions, findings and related information on an associated e-forum at http://www.evolvingcode.net/forum/viewforum.php?f=27.

Databases, Nucleic Acid↗
PubMed2006

An interactive visualization tool to explore the biophysical properties of amino acids and their contribution to substitution matrices.

BACKGROUND: Quantitative descriptions of amino acid similarity, expressed as probabilistic models of evolutionary interchangeability, are central to many mainstream bioinformatic procedures such as sequence alignment, homology searching, and protein structural prediction. Here we present a web-based, user-friendly analysis tool that allows any researcher to quickly and easily visualize relationships between these bioinformatic metrics and to explore their relationships to underlying indices of amino acid molecular descriptors. RESULTS: We demonstrate the three fundamental types of question that our software can address by taking as a specific example the connections between 49 measures of amino acid biophysical properties (e.g., size, charge and hydrophobicity), a generalized model of amino acid substitution (as represented by the PAM74-100 matrix), and the mutational distance that separates amino acids within the standard genetic code (i.e., the number of point mutations required for interconversion during protein evolution). We show that our software allows a user to recapture the insights from several key publications on these topics in just a few minutes. CONCLUSION: Our software facilitates rapid, interactive exploration of three interconnected topics: (i) the multidimensional molecular descriptors of the twenty proteinaceous amino acids, (ii) the correlation of these biophysical measurements with observed patterns of amino acid substitution, and (iii) the causal basis for differences between any two observed patterns of amino acid substitution. This software acts as an intuitive bioinformatic exploration tool that can guide more comprehensive statistical analyses relating to a diverse array of specific research questions.

Amino Acid Sequence↗
PubMed2006

Simplified gene synthesis: a one-step approach to PCR-based gene construction.

PCR-based gene synthesis conventionally requires two steps: first, all overlapping oligonucleotides are assembled by self-priming; then an additional pair of primers is used to amplify the full-length gene product. Here we propose a simplified method of gene synthesis which combines these two steps into one. We have found that the efficiency of this one-step method, which we term "Simplified Gene Synthesis", is affected by multiple parameters of the PCR reactions. In particular, the choice of polymerase is critical for successful one-step assembly. Other important factors include the concentration of assembly oligonucleotides and amplification primers. Moreover, we offer a general method to estimate, given a known mutation rate, how many clones should be sequenced in order to be confident of obtaining at least one correct gene product. Having determined the accuracy of gene products synthesized under optimal conditions with Simplified Gene Synthesis, we show that our estimation works well. Overall, the simplified gene synthesis provides an easier and more efficient approach to gene synthesis, providing a further step towards the future goal of generalized automation for this process.

Cloning, Molecular↗
PubMed2005

The Synthetic Gene Designer: a flexible web platform to explore sequence manipulation for heterologous expression.

"Codon optimization" is a general approach to improving heterologous expression where genes are moved from their native genomes into alternatives that exhibit different patterns of codon usage. However, despite reports of successful manipulations and the existence of stand-alone codon optimization software packages or commercial services that offer to redesign genes, the scientific community lacks any systematic understanding of what exactly it means to optimize codon usage. Thus we present a bona fide web application, the "Synthetic Gene Designer," which contrasts with existing software by providing a centralized, free, and transparent platform for the broader scientific community to develop knowledge about synthetic gene design. Consistent with this goal, our software is associated with a moderated e-forum that promotes discussion of synthetic gene design and offers technical support. In addition, the Synthetic Gene Designer presents enhanced functionality over existing software options: for example, it enables users to work with non-standard genetic codes, with user-defined patterns of codon usage and an expanded range of methods for codon optimization. The Synthetic Gene Designer, together with on-line tutorials and the forum, is available at .

Animals↗
PubMed2003

The case for an error minimizing standard genetic code.

Since discovering the pattern by which amino acids are assigned to codons within the standard genetic code, investigators have explored the idea that natural selection placed biochemically similar amino acids near to one another in coding space so as to minimize the impact of mutations and/or mistranslations. The analytical evidence to support this theory has grown in sophistication and strength over the years, and counterclaims questioning its plausibility and quantitative support have yet to transcend some significant weaknesses in their approach. These weaknesses are illustrated here by means of a simple simulation model for adaptive genetic code evolution. There remain ill explored facets of the 'error minimizing' code hypothesis, however, including the mechanism and pathway by which an adaptive pattern of codon assignments emerged, the extent to which natural selection created synonym redundancy, its role in shaping the amino acid and nucleotide languages, and even the correct interpretation of the adaptive codon assignment pattern: these represent fertile areas for future research.

Amino Acids↗