Towards Development of Robust Optimized Chassis (ROCs) for Whole Cell Modeling and Making Efficient, Safe, and Secure Biofactories
|演題||Towards Development of Robust Optimized Chassis (ROCs) for Whole Cell Modeling and Making Efficient, Safe, and Secure Biofactories|
|講演者||Prof. Barry Wanner（Purdue University, Biological Sciences）|
Abstract: A long-term goal of synthetic biology is the development of robust and optimized cells for whole cell modeling and production of innumerable bioproducts. Escherichia coli provides an excellent platform for engineering such cells because of our extensive knowledge about it and the vast number of tools for its facile manipulation. As a first step towards this goal, we have initiated an in silico design based on E. coli conserved proteins and conserved nucleotide contigs. The E. coli K-12 MG1655 model cell has 4666 protein-encoding genes, 14 16S and 23 S rRNA genes, 8 5S rRNA genes, 88 tRNA genes, a 4.5S and RNase P gene, and 60 other (mostly regulatory) non-coding RNAs (www.PATRICBRC.org). Comparisons of E. coli K-12, CFT072, and O157 genomes revealed a core genome of 2996 protein-coding genes in common (Welch et al., 2002). In more recent studies, comparison of nineteen E. coli genomes showed a core of (Rasko et al., 2008) protein-coding genes, and comparison of 61 E. coli genomes disclosed a core of 993 protein-coding genes (Lukjancenko et al., 2010). Our in silico design is based on the core proteome of 55 complete E. coli genomes and more one hundred complete or whole shotgun genomes (WSGs). We are in the process of integrating these genome and proteome datasets with manually and automated databases (ASAP, EcoCyc, EcoGene, PEC, RAST, and others) and transcriptome data from deep sequencing (RNA-seq) to identify functional regions important for growth on glucose minimal and rich media under conditions required for rapid growth, survival, and robustness. We plan to experimentally determine the function(s) of core genes and regions that are poorly described, and to use this information for development of a robust optimized cells by using technologies based on the lambda Red recombination method (Datsenko and Wanner, 2000) such as multiplex genome engineering and accelerated evolution (MAGE; (Wang et al., 2009)). Supported by NIH 1RC1GM092047
Datsenko,K.A. and Wanner,B.L. (2000). Proc. Natl. Acad. Sci. USA 97, 6640-6645.
Lukjancenko,O., Wassenaar,T.M., and Ussery,D.W. (2010). Microb. Ecol. 60, 708-720.
Rasko,D.A., Rosovitz,M.J., Myers,G.S., Mongodin,E.F., Fricke,W.F., Gajer,P., Crabtree,J., Sebaihia,M., Thomson,N.R., Chaudhuri,R., Henderson,I.R., Sperandio,V., and Ravel,J. (2008) J. Bacteriol. 190, 6881-6893.
Wang,H.H., Isaacs,F.J., Carr,P.A., Sun,Z.Z., Xu,G., Forest,C.R., and Church,G.M. (2009). Nature 460, 894-898.
Welch,R.A., Burland,V., Plunkett,G., III, Redford,P., Roesch,P., Rasko,D., Buckles,E.L., Liou,S.R., Boutin,A., Hackett,J., Stroud,D., Mayhew,G.F., Rose,D.J., Zhou,S., Schwartz,D.C., Perna,N.T., Mobley,H.L., Donnenberg,M.S., and Blattner,F.R. (2002). Proc. Natl. Acad. Sci. USA 99, 17020-17024.
森 浩禎 (firstname.lastname@example.org)