5.1 Microorganism, culture media, chemicals, BioLector and Software
Utilized the E. coli BL21 Rosetta 2 strain that had been transformed with the pET28a(+) vector containing the the modified insulin construct, featured a cleavable N-terminal 6XHis-tag. To assess the growth and expression of the the modified insulin clone, we employed Luria-Bertani (LB) Miller media as the foundational culture medium. The necessary chemical components were procured from reputable suppliers such as Merck and Sigma. Our internal resources provided a protein weight marker known as KUYBIIGM-Ladder. We utilized Design-Expert 7.0.0 software package (Stat-Ease, Inc., based in Minneapolis, MN, USA) to design and analyze optimization experiments. We employed the BioLector XT Microbioreactor (Brandd City, Country) to conduct screening experiments, considering statistically determined parameters.