InsectDirect™ System- pIEx™-6 DNA The pIEx™-6 vector is designed to allow rapid, high-level protein expression in insect cells without the time-consuming process of creating a recombinant baculovirus. This vector features the hr5 enhancer and the IE1 (immediate early) promoter to direct expression in insect cells. This promoter/enhancer combination recruits endogenous insect cell transcription machinery, thereby avoiding baculovirus infection and the associated cytotoxic effects. This plasmid features an amino-terminal His•Tag® coding sequence, and multiple cloning site (MCS) regions designed to allow the generation of target proteins with minimal vector-encoded fusion. The Pml I cloning site allows direct fusion to the His•Tag sequence for inserts that are blunt and in the appropriate reading frame. For applications requiring a removable amino-terminal His•Tag sequence, the MCS includes a PshA I cloning site (GACNNNNGTC). The PshA I site overlaps the cleavage site for the enterokinase (EK) protease (AspAspAspAspLys). Cloning appropriately designed inserts into this site re-creates the full EK site and allows all amino-terminal vector-encoded sequences to be removed by EK digestion. The remainder of the MCS encodes restriction enzyme sites found in many other Novagen expression vectors to facilitate insert transfer. An optional S•Tag™coding sequence is present at the distal end of the MCS for generating a carboxy-terminal tag compatible with purification, detection, and quantification (1). References:
1. Kim, J.S. and Raines, R.T. (1993) Protein Science 2, 348–356. |
