Figure 4.

Expression of functional BAF in Escherichia coli. (A) Overexpression, solubilization, and purification of the murine BAF. E. coli cells with the pET-15b recombinant plasmid containing BAF-ORF were harvested before (lane 1) or after (lanes 2–5) IPTG induction. Lanes: 1 and 2, whole cell lysates in 4% SDS; 3, soluble lysates; 4, guanidine hydrochloride-soluble portion of the insoluble fraction; 5, the purified BAF after removal of the His-tag and reverse-phase chromatography; 6, molecular mass standards. Samples were analyzed by SDS/PAGE, followed by Coomassie blue staining. (B) The recombinant BAF is active in the autointegration barrier reconstitution assay. Lanes: 1, no addition; 2–4, the purified BAF in 2, 10, and 50 ng in each reconstitution reaction.