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EGFP Translation Study Problem

EGFP Translation Study Problem - DNA Techniques

EGFP Translation Study Problem - Post questions and discuss DNA techniques and protocols such as DNA extraction, PCR, and the study of DNA-binding proteins.


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Old 01-15-2009, 12:49 AM
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Default EGFP Translation Study Problem



Hi all!
I am performing a translation study in mammalian cell culture. My construct includes: universal promoter of the vector, leader sequence of my gene of interest (which has a meaning in a translation efficiency), EGFP and FLAG tag. Using antibodies against FLAg I am able to obtain band of fusion protein of expected size in western blot - therefore I concluded that translation takes place in my culture. But EGFP signal is absent, when cells are observed under the fluorescent microscope.
Any ideas why I am having this problem?
Thanks
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Old 10-14-2011, 04:23 PM
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Default Re: EGFP Translation Study Problem

based on ur observation it is highly possible that your fusion protein (GeneX-EGFP-FLAG) does not fluoresce. EGFP is quite sensitive, any excess amount of residues fused to it will drastically alter its folding properties, hence its fluorescence intensity.
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