i am trying to express a gene in bacterial cells but i am unable to express it. i have done the codon optimisation and have thoroughly checked the sequence as well. i have also tried various strains like bl21de3, c41, rosetta, plysS gold, t7 and have also tried growth media like super and terrific broth yet could not suceed. please help me out.
You should select appropriate promoter.I think it would be helpful to express the protein of interest.
Dear Soumya
the reasons because a gene can not show any expression in E.coli are several and can change on the basis of the gene properties. ( protein MW, presence of signal peptides, menbrane region, isoelettric point, domain desing)
Can you provide to me some more detail about gene suquence and the Vectors that you used for cloning and expression?
Hi,
Besides of useful comments above, we need an info about your expression plasmid. You should use proper expression plasmid with proper bacterial strain.
Maybe your protein of interest was degraded during long cultivation. Or induction agent concentration (IPTG, I assume) was too high and bacteria grew too fast. So the bacteria did not have enough time to complete the protein.
If you give a little more information about your protocol, it will be useful to solve your problem.
Good luck
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