pET-28a-chBCO2

Protocol tips

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	For chicken BCO2, the plasmid was transformed into E. coli BL21-Gold (DE3). The transformed bacterial cells were grown in LB broth (Sigma-Aldrich) at 30 °C to an A600 of 0.5–0.7. The incubator temperature was then lowered to 16 °C, and expression of the recombinant protein was induced by adding isopropyl 1-thio-β-d-galactopyranoside (Gold Biotechnology) to a final concentration of 0.1 mm.

Protocol tips
For chicken BCO2, the plasmid was transformed into E. coli BL21-Gold (DE3). The transformed bacterial cells were grown in LB broth (Sigma-Aldrich) at 30 °C to an A600 of 0.5–0.7. The incubator temperature was then lowered to 16 °C, and expression of the recombinant protein was induced by adding isopropyl 1-thio-β-d-galactopyranoside (Gold Biotechnology) to a final concentration of 0.1 mm.

Publication protocol

For chicken BCO2, the plasmid was transformed into E. coli BL21-Gold (DE3). The transformed bacterial cells were grown in LB broth (Sigma-Aldrich) at 30 °C to an A600 of 0.5–0.7. The incubator temperature was then lowered to 16 °C, and expression of the recombinant protein was induced by adding isopropyl 1-thio-β-d-galactopyranoside (Gold Biotechnology) to a final concentration of 0.1 mm. Five liters of culture were grown for 16 h, and the cells were harvested by centrifugation. The cell pastes were frozen at −80 °C and thawed prior to lysis. The protein was purified using the same protocol for recombinant human BCO1 using cobalt ion affinity chromatography as in our previous publication (9).

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