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Analysis of shuffled gene libraries

Joern, John M and Meinhold, Peter and Arnold, Frances H. (2002) Analysis of shuffled gene libraries. Journal of Molecular Biology, 316 (3). pp. 643-656. ISSN 0022-2836. doi:10.1006/jmbi.2001.5349. https://resolver.caltech.edu/CaltechAUTHORS:20170408-161123738

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Abstract

In vitro recombination of homologous genes (family shuffling) has been proposed as an effective search strategy for laboratory evolution of genes and proteins. Few data are available, however, on the composition of shuffled gene libraries, from which one could assess the efficiency of recombination and optimize protocols. Here, probe hybridization is used in a macroarray format to analyze chimeric DNA libraries created by DNA shuffling. Characterization of hundreds of shuffled genes encoding dioxygenases has elucidated important biases in the shuffling reaction. As expected, crossovers are favored in regions of high sequence identity. A sequence-based model of homologous recombination that captures this observed bias was formulated using the experimental results. The chimeric genes were found to show biases in the incorporation of sequences from certain parents, even before selection. Statistically different patterns of parental incorporation in genes expressing functional proteins can help to identify key sequence-function relationships.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1006/jmbi.2001.5349DOIArticle
ORCID:
AuthorORCID
Arnold, Frances H.0000-0002-4027-364X
Additional Information:© 2002 Elsevier. Received 5 October 2001, Revised 14 December 2001, Accepted 14 December 2001, Available online 5 March 2002. The authors thank Jonathan Silberg, Christopher Voigt and Lillian Pierce for their comments on the manuscript. This work was supported by the U. S. National Science Foundation (BES#9981770) and by Maxygen, Inc.
Funders:
Funding AgencyGrant Number
NSFBES-9981770
Maxygen, Inc.UNSPECIFIED
Subject Keywords:directed evolutionrecombinationfamily shufflingStEPrandom chimeragenesis
Issue or Number:3
DOI:10.1006/jmbi.2001.5349
Record Number:CaltechAUTHORS:20170408-161123738
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20170408-161123738
Official Citation:John M. Joern, Peter Meinhold, Frances H. Arnold, Analysis of shuffled gene libraries , Journal of Molecular Biology, Volume 316, Issue 3, 2002, Pages 643-656, ISSN 0022-2836, http://dx.doi.org/10.1006/jmbi.2001.5349. (http://www.sciencedirect.com/science/article/pii/S0022283601953495)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:76143
Collection:CaltechAUTHORS
Deposited By: 1Science Import
Deposited On:22 Jun 2017 20:38
Last Modified:15 Nov 2021 16:57

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