The theory is that it's using the lac operon which is normally turned off using a supressor. Once you add IPTG which is nonhydrolyzable but similar to lactose the activator, it binds and removes the suppressor. But because it can't be hydrolyzed, the protein expression doesn't get turned off by a feedback loop and stays on. U also need a good number of bacteria (OD 0.7 is ok). Too low you get low expression, too high they die. Lmk if u need a seperate video.
I have a question. Suppose I have a handful of different proteins and on the other hand different cells are present. How to design an experiment to know which protein is expressed in which cell?
@@Ashishkumar-id1nn So when you order the plasmid for a protein you know what it is and you transform it into a known cell. You have to label it and during the entire expression you must label everything so you don't mix up the different protein and cells. I don't recommend expressing them on the same day because of the risk to mix them up unless you really have to. If you mix them up then u can use mass spec to identify which is which. That would be a waste of money if you could label everything and keep track of it.
@@Ashishkumar-id1nn If you question is something else and you mean everything is unknown from the beginning, the best methods would be some kind of fluorescence assay and microscopy. Detection of mRNA levels are good but don't always mean they get translated.
