Three genes amplified from start codon ATG to stop codon TAG. If I fused them into one single fragment using overlapping PCR and cloned them into a vector under a single promter, could I get three indepedent proteins or could all three genes successfully express in bacteria?
In generale, it was very hard. I suggested that these genes should be inserted into to seperated promoter-guided location of the vector, or you can try to use a bidirectional promoter but still express only two genes at the same time.
This is a really bad idea. Put each gene under a promotor. Even if its the same promotor is ok. But one for all of the is not going to work at all. An other thing is that you can put a linker between the proteins and in this linker introduce the sequence for TEV protease cleavage.. Then you will have a protein fusion that you can cleave. However the best thing is ti clone in different promoters or express independently. Unless, it is necessary to express in the same cell. If this is the case, I would go for cloning each one under one promoter.
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