pMCSG7-D1aerobic

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	For expression under aerobic conditions BL21 (DE3)RILP cells harboring the pMCSG7 plasmid with the D1 gene were grown in 50 mL of Lennox LB medium (#LBL405, BioShop Canada Inc.) with 100 μg/mL ampicillin (#K029.4, Carl Roth) and 34 μg/mL chloramphenicol (#227920250, Acros Organics) overnight at 37 °C with shaking. The next day, the culture was transferred to 4 L of Terrific-Broth (TB) medium (#TER409, BioShop Canada Inc.) with ampicillin and chloramphenicol for selection, and cells were grown at 37 °C until the culture reached an OD600 of 1.2. The temperature was decreased to 18 °C, and cells were induced with 0.15 mM isopropyl β-d-1-thiogalactopyranoside (IPTG) (#CN08.3, Carl Roth).

Protocol tips

For expression under aerobic conditions BL21 (DE3)RILP cells harboring the pMCSG7 plasmid with the D1 gene were grown in 50 mL of Lennox LB medium (#LBL405, BioShop Canada Inc.) with 100 μg/mL ampicillin (#K029.4, Carl Roth) and 34 μg/mL chloramphenicol (#227920250, Acros Organics) overnight at 37 °C with shaking. The next day, the culture was transferred to 4 L of Terrific-Broth (TB) medium (#TER409, BioShop Canada Inc.) with ampicillin and chloramphenicol for selection, and cells were grown at 37 °C until the culture reached an OD600 of 1.2. The temperature was decreased to 18 °C, and cells were induced with 0.15 mM isopropyl β-d-1-thiogalactopyranoside (IPTG) (#CN08.3, Carl Roth).

Publication protocol

For expression under aerobic conditions BL21 (DE3)RILP cells harboring the pMCSG7 plasmid with the D1 gene were grown in 50 mL of Lennox LB medium (#LBL405, BioShop Canada Inc.) with 100 μg/mL ampicillin (#K029.4, Carl Roth) and 34 μg/mL chloramphenicol (#227920250, Acros Organics) overnight at 37 °C with shaking. The next day, the culture was transferred to 4 L of Terrific-Broth (TB) medium (#TER409, BioShop Canada Inc.) with ampicillin and chloramphenicol for selection, and cells were grown at 37 °C until the culture reached an OD600 of 1.2. The temperature was decreased to 18 °C, and cells were induced with 0.15 mM isopropyl β-d-1-thiogalactopyranoside (IPTG) (#CN08.3, Carl Roth). Protein production took place overnight at 18 °C. Cells were centrifuged and the pellet was stored at −80 °C for a few days before purification.

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