alexa Coupling mutagenesis and parallel deep sequencing to probe essential residues in a genome or gene.
Bioinformatics & Systems Biology

Bioinformatics & Systems Biology

Journal of Next Generation Sequencing & Applications

Author(s): Robins WP, Faruque SM, Mekalanos JJ

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Abstract The sequence of a protein determines its function by influencing its folding, structure, and activity. Similarly, the most conserved residues of orthologous and paralogous proteins likely define those most important. The detection of important or essential residues is not always apparent via sequence alignments because these are limited by the depth of any given gene's phylogeny, as well as specificities that relate to each protein's unique biological origin. Thus, there is a need for robust and comprehensive ways of evaluating the importance of specific amino acid residues of proteins of known or unknown function. Here we describe an approach called Mut-seq, which allows the identification of virtually all of the essential residues present in a whole genome through the application of limited chemical mutagenesis, selection for function, and deep parallel genomic sequencing. Here we have applied this method to T7 bacteriophage and T7-like virus JSF7 of Vibrio cholerae.
This article was published in Proc Natl Acad Sci U S A and referenced in Journal of Next Generation Sequencing & Applications

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