To transfect cells with a plasmid coding for Renilla luciferase (Rluc) in addition to the stable cell line expressing the GPCR of interest with a YFP tag, you can use a transfection reagent such as Lipofectamine or PEI.
Here are some general steps to follow:
It's important to optimize the transfection conditions such as the amount of DNA and transfection reagent used, as well as the incubation time, to achieve the best transfection efficiency and minimal toxicity.
Thanks a lot,
what about doxycycline? Should it be added before or after transfection with Rluc?
Doxycycline is an antibiotic that is commonly used in research to regulate gene expression in cells that have been genetically modified to express inducible systems. The timing of doxycycline addition will depend on the specific experimental design and the goal of the experiment.
If the goal of the experiment is to induce gene expression after transfection with Rluc, doxycycline can be added after transfection to initiate the expression of the inducible system. However, if the goal is to inhibit gene expression, doxycycline can be added before transfection to suppress the expression of the inducible system.
It is important to note that the optimal timing and concentration of doxycycline may vary depending on the specific cell type and inducible system being used. Therefore, it is recommended to optimize the experimental conditions for each specific application.
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