Hey guys,
I am currently cloning enzymes from a marine Lenthimonas-related isolate into E.coli NEBalpha for transformation of my vector (pet28a) and E.coli BL21 for overexpression of the target enzyme. My sulfatases are expressed but inactive and I have red that some sulfatases need co-expression with FGE enzymes in order to activate them.
I was thinking of transforming two different vectors (one for the sulfatases and one for FGE) with different antibiotic resistance into E.coli BL21.
Do I have to consider anything specific for my sulfatases (or FGE) regarding primer design or setting up the expression system?
Thanks in advance!
Two options:
1. Use an expression plasmid that can express multiple proteins, e.g. pET-Duet (there are others)
2. Use 2 plasmids, but then make sure to have to different compatible origins (e.g. p15A and pBR/pUC) and 2 different antibiotic selection markers to assure the two plasmids are indeed present.
Also see the discussion here
https://www.researchgate.net/post/Which_plasmid_to_use_for_expression_of_two_proteins
Yoram
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