Skip Navigation

Nucleic Acids Research 2006 34(4):e30; doi:10.1093/nar/gnj032
This Article
Right arrow Full Text Freely available
Right arrow Print PDF (997K) Freely available
Right arrow Screen PDF (995K) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Commercial Re-use Guidelines
for Open Access NAR Content
Google Scholar
Right arrow Articles by Fujii, R.
Right arrow Articles by Hayashi, K.
Right arrow Search for Related Content
PubMed
Right arrow Articles by Fujii, R.
Right arrow Articles by Hayashi, K.
Related Collections
Right arrow Mutagenesis
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

Published online 21 February 2006

© The Author 2006. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oxfordjournals.org

Methods Online

RAISE: a simple and novel method of generating random insertion and deletion mutations

Ryota Fujii, Motomitsu Kitaoka* and Kiyoshi Hayashi

National Food Research Institute 2-1-12 Kannondai, Tsukuba, Ibaraki, 305-8642, Japan

*To whom correspondence should be addressed. Tel: +81 29 838 8071; Fax: +81 29 838 7321; Email: mkitaoka{at}affrc.go.jp

Received November 4, 2005. Revised January 24, 2006. Accepted February 4, 2006.

Although proteins may be artificially improved by random insertion and deletion mutagenesis methods, these procedures are technically difficult, and the mutations introduced are no more variable than those introduced by the introduction of random point mutations. We describe here a three-step method called RAISE, which is based on gene shuffling and can introduce a wide variety of insertions, deletions and substitutions. To test the efficacy of this method, we used it to mutate TEM ß-lactamase to generate improved antibiotic resistance. Some unique insertion or deletion mutations were observed in the improved mutants, some of which caused higher activities than point mutations. Our findings indicate that the RAISE method can yield unique mutants and may be a powerful technique of protein engineering.

Present address: Ryota Fujii, Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, 1479 Gortner Avenue, Saint Paul, MN 55108, USA


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 Protein Eng Des SelHome page
S. Emond, P. Mondon, S. Pizzut-Serin, L. Douchy, F. Crozet, K. Bouayadi, H. Kharrat, G. Potocki-Veronese, P. Monsan, and M. Remaud-Simeon
A novel random mutagenesis approach using human mutagenic DNA polymerases to generate enzyme variant libraries
Protein Eng. Des. Sel., April 1, 2008; 21(4): 267 - 274.
[Abstract] [Full Text] [PDF]

Home page
 Protein Eng Des SelHome page
A. Herman and D. S. Tawfik
Incorporating Synthetic Oligonucleotides via Gene Reassembly (ISOR): a versatile tool for generating targeted libraries
Protein Eng. Des. Sel., May 5, 2007; (2007) gzm014v1.
[Abstract] [Full Text] [PDF]


Disclaimer: Please note that abstracts for content published before 1996 were created through digital scanning and may therefore not exactly replicate the text of the original print issues. All efforts have been made to ensure accuracy, but the Publisher will not be held responsible for any remaining inaccuracies. If you require any further clarification, please contact our Customer Services Department.