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Why is the TurboGFP expression level low for my Precision LentiORF construct?
It is because the TurboGFP is being expressed after an IRES. In our hands IRES-TurboGFP has always produced lower protein expression level compared to the upstream ORF.
In addition, in pLOC, TurboGFP is expressed in form of a fusion TurboGFP-2A-BlastR. This fusion marker is translated into TurboGFP-2A and BlastR. The residual 2A peptide at 3' of TurboGFP also affects GFP brightness quite a bit. This was a reason why we decided to direct GFP to the nucleus - to concentrate the signal as much as possible.