From Gateway to MultiSite Gateway in one recombination event.
BACKGROUND
Invitrogen Gateway technology exploits the integrase/att site-specific recombination system for directional cloning of PCR products and the subsequent subcloning into destination vectors. One or three DNA segments can be cloned using Gateway or MultiSite Gateway respectively. A vast number of single-site Gateway destination vectors have been created while MultiSite Gateway is limited to few destination vectors and therefore to few applications. The aim of this work was to make the MultiSite Gateway technology available for multiple biological purposes.


RESULTS
We created a construct, pDONR-R4-R3, to easily convert any available Gateway destination vector to a MultiSite Gateway vector in a single recombination reaction. In addition, we designed pDONR-R4-R3 so that DNA fragments already cloned upstream or downstream of the Gateway cassette in the original destination vectors can still be utilized for promoter-gene or translational fusions after the conversion.


CONCLUSIONS
Our tool makes MultiSite Gateway a more widely accessible technology and expands its applications by exploiting all the features of the Gateway vectors already available.
Magnani
Enrico
E
Department of Plant and Microbial Biology, University of California, Berkeley, California 94720, USA. emagnani@berkeley.edu
Bartling
Linnea
L
Hake
Sarah
S
D003001:Cloning, Molecular; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000378:metabolism; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005821:Genetic Techniques; D005822:Genetic Vectors; D016678:Genome; D019426:Integrases / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011995:Recombination, Genetic
D004274:DNA, Recombinant; D019426:Integrases