Site-specific recombination in Schizosaccharomyces pombe and systematic assembly of a 400kb transgene array in mammalian cells using the integrase of Streptomyces phage phiBT1.
We have established the integrase of the Streptomyces phage phiBT1 as a tool for eukaryotic genome manipulation. We show that the phiBT1 integrase promotes efficient reciprocal and conservative site-specific recombination in vertebrate cells and in Schizosaccharomyces pombe, thus establishing the utility of this protein for genome manipulation in a wide range of eukaryotes. We show that the phiBT1 integrase can be used in conjunction with Cre recombinase to promote the iterative integration of transgenic DNA. We describe five cycles of iterative integration of a candidate mouse centromeric sequence 80 kb in length into a human mini-chromosome within a human-Chinese hamster hybrid cell line. These results establish the generality of the iterative site-specific integration technique.
Xu
Zhengyao
Z
Institute of Genetics, Queen's Medical Centre, University of Nottingham, Nottingham NG7 2UH, UK.
Lee
Nicholas C O
NC
Dafhnis-Calas
Felix
F
Malla
Sunir
S
Smith
Margaret C M
MC
Brown
William R A
WR
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D002645:Chickens; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D019426:Integrases / Q000378:metabolism*; D051379:Mice; D011995:Recombination, Genetic; D012568:Schizosaccharomyces / Q000235:genetics*; D017906:Siphoviridae / Q000201:enzymology*; D013302:Streptomyces / Q000821:virology; D019076:Transgenes
C045073:Cre recombinase; D019426:Integrases