Figure 5

Impact of the temperature on Mos1 transposition. (a) Effect of the temperature (horizontal axis) on the transposition efficiency (vertical axis). In vitro assays were performed using two plasmids (pET-3Tet3 as the transposon donor, and pBC-SK+ as the target for integration) and purified MBP-MOS1. Two different shapes of pBC-3Tet3 were used: the super-helical form (gray bars) and that linearized with Nco I (empty bars). (b) Effect of temperature on the excision. Time course analyses were done using super coiled pBC-3Tet3 as pseudo-Mos1 and purified MBP-MOS1. The assays were performed at 30°C (top panel) or 37°C (bottom panel) and the resulting products were loaded onto BET-stained agarose gel. Molecular weight markers are indicated in the left margin. The various products are depicted on the right and their positions on the gel are indicated. OC: open circle; Lin: linear; SC: super coiled; 3Tet3: excised transposon. On the top panel (30°C) the expected position of 3Tet3 is pointed out, but the corresponding product is not detected. Asterisks indicate the integration products, determined according to [38] and personal data.