File:BE).png

D、IPTG(-/+) = without/with 0.2 mM IPTG for 3-6 hours, adding IPTG to a bacteria culture with OD600 0.2-0.3. M: Protein molecular weight marker ladder. Lane 1~4、6～15: pET28 plasmids encoding crtIYBE separated by self-cleaving ribozyme（BBa_K4162021），crtI+crtY+crtB+crtE，crtI+crtY，crtE+crtI，crtB+crtE without any tag were transformed into BL21(DE3) HI-Control Rosetta strain, single clones (6a, 4a, IY10, EI8，BE11，BE12，BE13)were picked for liquid LB culture. 6=crtIYEB，4=crtIYBE. Protein expression was induced in parallel cultures by IPTG. Bacterial cultures were monitored by OD600, and 5x10^7 cells were harvested by centrifugation and lysis in 1x SDS sample buffer. Equal amount (10 μL, 2x10^6 cells) of whole cell lysate were analyzed by SDS-PAGE (14% separation gel). Red arrows point to crtI protein. Green arrows point to crtY protein. Black arrows point to crtB protein. Yellow arrows point to crtE protein.