Nucleic Acids Research, 2002, Vol. 30, No. 11 2299-2306
© 2002 Oxford University Press
Enhanced efficiency through nuclear localization signal fusion on phage 
C31-integrase: activity comparison with Cre and FLPe recombinase in mammalian cells
Artemis Pharmaceuticals GmbH, Neurather Ring 1, 51063 Köln, Germany
The integrase of the phage 
C31 recombines an attP site in the phage genome with a chromosomal attB site of its Streptomyces host. We have utilized the integrase-mediated reaction to achieve episomal and genomic deletion of a reporter gene in mammalian cells, and provide the first comparison of its efficiency with other recombinases in a new assay system. This assay demonstrated that the efficiency of C31-integrase is significantly enhanced by the C-terminal, but not the N-terminal, addition of a nuclear localization signal and becomes comparable with that of the widely used Cre/loxP system. Furthermore, we found that the improved FLP recombinase, FLPe, exhibits only 10% recombination activity on chromosomal targets as compared with Cre, whereas the Anabaena derived XisA recombinase is essentially inactive in mammalian cells. These results provide the first demonstration that a nuclear localisation signal and its position within a recombinase can be important for its efficiency in mammalian cells and establish the improved C31-integrase as a new tool for genome engineering.
* To whom correspondence should be addressed. Tel: +49 221 9645310; Fax: +49 221 9645321; Email: r.kuehn{at}artemispharma.de
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  E. Sangiorgi, Z. Shuhua, and M. R. Capecchi In vivo evaluation of PhiC31 recombinase activity using a self-excision cassette Nucleic Acids Res., November 1, 2008; 36(20): e134 - e134. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Z. Xu, N. C. O. Lee, F. Dafhnis-Calas, S. Malla, M. C. M. Smith, and W. R. A. Brown Site-specific recombination in Schizosaccharomyces pombe and systematic assembly of a 400kb transgene array in mammalian cells using the integrase of Streptomyces phage {phi}BT1 Nucleic Acids Res., January 17, 2008; 36(1): e9 - e9. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Hitz, W. Wurst, and R. Kuhn Conditional brain-specific knockdown of MAPK using Cre/loxP regulated RNA interference Nucleic Acids Res., June 22, 2007; (2007) gkm475v1. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Gupta, R. Till, and M. C. M. Smith Sequences in attB that affect the ability of {phi}C31 integrase to synapse and to activate DNA cleavage Nucleic Acids Res., May 11, 2007; 35(10): 3407 - 3419. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Bischof, R. K. Maeda, M. Hediger, F. Karch, and K. Basler An optimized transgenesis system for Drosophila using germ-line-specific {varphi}C31 integrases PNAS, February 27, 2007; 104(9): 3312 - 3317. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J.-z. Chen, C.-n. Ji, G.-l. Xu, R.-y. Pang, J.-h. Yao, H.-z. Zhu, J.-l. Xue, and W. Jia DAXX interacts with phage {Phi}C31 integrase and inhibits recombination Nucleic Acids Res., December 4, 2006; 34(21): 6298 - 6304. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. K. Moore and V. Srivastava Efficient Deletion of Transgenic DNA from Complex Integration Locus of Rice Mediated by Cre/lox Recombination System Crop Sci., February 1, 2006; 46(2): 700 - 705. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Dafhnis-Calas, Z. Xu, S. Haines, S. K. Malla, M. C. M. Smith, and W. R. A. Brown Iterative in vivo assembly of large and complex transgenes by combining the activities of {varphi}C31 integrase and Cre recombinase Nucleic Acids Res., December 15, 2005; 33(22): e189 - e189. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Chen and S. L. C. Woo Complete and persistent phenotypic correction of phenylketonuria in mice by site-specific genome integration of murine phenylalanine hydroxylase cDNA PNAS, October 25, 2005; 102(43): 15581 - 15586. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. T. Wong, J. L Kolman, Y.-C. Li, L. D Mesner, W. Hillen, C. Berens, and G. M Wahl Reproducible doxycycline-inducible transgene expression at specific loci generated by Cre-recombinase mediated cassette exchange Nucleic Acids Res., October 4, 2005; 33(17): e147 - e147. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Marillonnet, A. Giritch, M. Gils, R. Kandzia, V. Klimyuk, and Y. Gleba In planta engineering of viral RNA replicons: Efficient assembly by recombination of DNA modules delivered by Agrobacterium PNAS, May 4, 2004; 101(18): 6852 - 6857. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Lauth, F. Spreafico, K. Dethleffsen, and M. Meyer Stable and efficient cassette exchange under non-selectable conditions by combined use of two site-specific recombinases Nucleic Acids Res., November 1, 2002; 30(21): e115 - e115. [Abstract] [Full Text] [PDF]
|
|