Restriction digests of the clone give the following sizes (kb): EcoRI--1.3; HindIII--1.3; BglII--1.3. The vector contains the following restriction sites (approximate kb from nt 1): BglII--0.79; ClaI--0.11; EcoRI--0.33; HindIII--0.02; NcoI--0.12; PstI--0.75; PvuII--0.23. Replication depends on read-through from the tac promoter. Replication may be negatively affected by cloning of large inserts or sequences containing a transcriptional terminator. Expression vector permitting production of heterologous peptides fused to active chloramphenicol acetyltransferase. Repression of this high copy number plasmid is hard to achieve, even using hosts overproducing lacIq. Escherichia coli WK6 is recommended for inducible expression. The replicon is derived from ColE1 and does not contain the RNAII promoter nor most of the antisense RNAI control element coding sequence. This allows compatibility with other ColE1 plasmids. The order of the major features of the plasmid is: HindIII - tac promoter - NcoI - cat/ScaI/MCS/BglII - HyRep2 ori. |
