All our multiexpression systems (currently MultiBac, MultiMam and MultiColi) rely on in vitro Cre-loxP recombination of Acceptor and Donor plasmids to assemble multiprotein expression vectors. A theoretically unlimited number of plasmids can be fused in this way, currently the systems (pragmatically) foresee up to three Donor plasmids to be recombined with a single Acceptor plasmid to yield the product plasmid which is the multigene expression construct(s) of choice.

Each educt plasmid (Donors or Acceptor) contains a unique resistance marker to allow selection of productive fusion plasmids by multiple resistance marker challenge. This approach is made possible by the conditional origin of replication present on all Donors which requires their fusion to an Acceptor to allow for propagation in a regular bacterial host strain which does not recognize the conditional origin. The product plasmid is barcoded by the resistance markers present on the Acceptor and Donor plasmids used in the Cre-LoxP fusion reaction, this is the basis for selection by using appropriate combinations of antibiotics.

Cre-ACEMBLER was programmed by Christian Becke, now at Freie Universitat Berlin.

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