Recombination system based on cre alpha complementation and leucine zipper fusions.
In a previous study, a recombination system based on the alpha complementation of cre recombinase and protein transduction was established. This system relied on the transient expression of the inactive, self-excisable C-terminal (beta) and the transduction of the N-terminal (alpha) cre fragments to cells as a purified protein. This recombination system potentially results in a less invasive and more controllable cre recombination in mammalian cells. In this study, we have employed a more efficient complementation triggering sequence using more than only the overlapping amino acids to help the alpha and beta fragments reassociate. In order to increase the association efficiency of the complementing fragments of cre recombinase, we chose to use a fusion of cre fragments to a self-heterodimerizing pair of proteins to trigger their binding and thus increase the efficiency of the restored enzymatic activity. For this purpose, the leucine zipper motifs (bJun and bFos) of the AP-1 transcription were fused to cre fragments (alpha and beta, respectively). This resulted in an increased reassociation efficiency of the fragments and a two times more efficient recombination system compared with the previous study.
Seidi
Azadeh
A
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501, Japan.
Mie
Masayasu
M
Kobatake
Eiry
E
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D019426:Integrases / Q000235:genetics / Q000378:metabolism*; D016350:Leucine Zippers / Q000235:genetics* / Q000502:physiology; D008954:Models, Biological; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic; D018808:Transcription Factor AP-1 / Q000235:genetics / Q000378:metabolism; D014161:Transduction, Genetic
D011993:Recombinant Fusion Proteins; D018808:Transcription Factor AP-1; C045073:Cre recombinase; D019426:Integrases