The "2in1" cloning system

Analysing higher order protein complexes requires introduction of multiple plasmids into a host organism. This becomes cumbersome and to a certain extent unreliable as it cannot be guaranteed that all plasmids will be transformed into a single cell. We have devised a novel, Gateway-compatible cloning system - 2in1 - that enables single-step transfer of two independent genes into two different expression cassettes on the same vector backbone (Grefen and Blatt 2012b).

This 2in1 system can be readily transferred in other cell and expression systems (Hecker et al. 2015). We have created a set of helper vectors similar to Life Technologies’ Gateway conversion kit that allows conversion of any expression vector providing high flexibility for the researcher.

The use of the 2in1 cloning system is not limited to improving protein-protein interaction methods; this concept offers advantages for all techniques that depend on multiple gene expression, by generally circumventing issues inherent to work with individually cloned genes (Karnik et al. 2013b). It allows increasing the number of genes to be introduced in a foreign organism as it decreases the number of plasmids, and thereby auxotrophies and other selection markers, necessary for the purpose.