Post Lab Questions | Mandatory for All Students

1. Use the images obtained in the lab or the ones attached and quantify fluorescence fold change between treatments. Discuss the functionality of the lacI protein in reference to this experiment a the genetic circuit used.

From ChatGPT: LacI is a repressor protein that binds to the operator region of the lac operon, preventing transcription in the absence of an inducer (like IPTG). In this experiment, the genetic circuit includes a reporter gene (e.g., GFP) under the control of a LacI-regulated promoter. In the absence of IPTG (0X), LacI binds to the operator, and GFP expression is repressed (no fluorescence). As IPTG concentration increases, IPTG binds LacI, causing a conformational change that prevents LacI from binding to the operator. This de-represses the promoter, allowing transcription of GFP → increased fluorescence.

1. What we should expect if the induction was made inside of a bacteria like E.coli BL21 DE3?

From ChatGPT: If we move this IPTG-inducible LacI-GFP system into E. coli BL21(DE3): We expect similar induction patterns but with slower kinetics. Fluorescence will increase with IPTG, but onset is delayed due to cellular processes. Expression might be stronger, especially if using a T7-based system. Cellular burden and regulation (e.g., plasmid copy number, proteolysis) can influence the results.