α lactose agarose

MGC human galectin-1–encoding cDNA (LGALS1, accession# BC001693) was PCR-amplified with the CACC forward primer (FWD: 5′-CAC CAT GGC TTG TGG TCT GG-3′) and reverse primer (REV: 5′-TCA GTC AAA GGC CAC ACA TTT GAT CT-3′). The amplified product was subcloned into pET101 by directional TOPO expression. The cloned product was transformed into One SHOT TOP10 Escherichia coli by heat-shock. Cells were grown overnight on ampicillin. Resistant cultures were selected and were used for DNA isolation. BL21 StarTM One Shot cells were transformed with the pET101 vector with LGALS1 by heat-shock, brief outgrowth in SOC medium, followed by transfer to 10 ml of LB containing ampicillin, and the cultures were grown overnight at 37°C while shaking. The next day, 50 ml of LB containing ampicillin was inoculated with 1 ml of the overnight culture. The culture was grown at 37°C with shaking (225 to 250 rpm) for 2 to 3 hours. IPTG (1 mM) was added to induce the expression of galectin-1 for 3 to 4 hours. The cells were then harvested by centrifugation at 3000g for 10 min at 4°C. The cells were purified by α-lactose/agarose (Sigma-Aldrich) as detailed previously (88 (link)).