pET16b plasmid transformation

Fifty microlitres of competent E. coli BL21 (DE3) cells (New England BioLabs) were combined with 5 ng of pET16b plasmid DNA (Addgene) and incubated on ice for 30 min. The pET16b plasmid contained the ampR gene and promoter for inducible expression of the E. coli chromosomal ampC gene. Contents were subjected to heat shock for 10 s at 42°C followed by a 5 min incubation on ice. Nine hundred and fifty microlitres of super optimal broth with catabolite repression media were added, and the transformed cells were incubated at 37°C with 180 rpm shaking for 1 h. The resulting culture was spread onto Luria‐Bertani (LB) agar plates containing 50 μg ml⁻¹ ampicillin sodium (Formedium) and incubated overnight at 37°C for plasmid uptake selection.