A novel, single plasmid, approach to in-frame fusion

The sequence GCGCGCGCGC contains three overlapping, mutually exclusive, BssHII restriction sites, each corresponding to one of the three different reading frames. When inserted into a plasmid and digested with BssHII, the three sites in this sequence are cleaved at an approximate ratio of 2:1:2. Consequently, this system can be used to simplify in vitro in frame fusions to involve a single plasmid. We have constructed such a plasmid and used it to select an open reading frame in a 1.6 kb cDNA fragment.