pMXB10 is a control plasmid for the IMPACT™ Kit (NEB #E6901) (1,2). This plasmid carries the E. coli malE gene, encoding the maltose binding protein (MBP, 42 kDa) (3), fused in-frame to the coding region of the Mxe gyrase A intein-chitin binding domain (28 kDa) from pTXB1 (2,4,5). pMXB10 can be used to test plasmid transformation, cell culture, induction and purification procedures. After induction with 0.3 mM IPTG at 30°C for 3 hours (or 15°C for 12-16 hours), 100 ml of cells should yield 2-3 mg of a 70 kDa fusion protein. After chitin column purification and cleavage, approximately 1.5-2.0 mg of MBP is usually obtained. This double stranded vector is 7813 bp in length.
- Expression of the fusion gene is under the control of an IPTG -inducible T7 promoter (5).
- T7 Express Competent E. coli (High Efficiency) (NEB #C2566) or BL21(DE3) Competent E. coli (NEB #C2527) and derivatives can be used as expression hosts.
- A pTXB1 derivative
- Ampicillin resistance
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