I have tagged my interested protien with GFP to observe the localization one of them can be perfectly observed but another 2 did not? I wonder that what's wrong with them since there is no problem about DNA sequences.
Hi Natthaporn,
Do you have any linkers between your protein and GFP? If so, make sure that there is no frame shift.
Can you detect expression of the fusion protein in the strains with an anti-GFP antibody on a Western blot? That would tell you if the fusion protein is being expressed at all, and if the protein is the predicted size.
Are you talking about 3 different proteins, or a single protein with 3 different transformation events?
I mean 3 different protein tagged with GFP. One is native, one is its homolog and one which added with MTS but basically these 3 protein are very similar. I also have a linker and I have checked all of the whole sequences, there is no frameshift.
By MTS, you meant Mitochondrial Targeting Sequence? And which one of them you saw the GFP fluorescence? The one with MTS? Was it successfully targeted to mitochondrial? Did you have mitochondria marker to verify that?
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