Hello Yogi,

Most of the time, human proteins in bacteria is difficult to produce due to the host issues; we mainly encounter a little yield or improper folding/inclusion bodies.You really have to spend your time to establish a nice way on this primary issues. I feel a solubility enhancing tag is always helpful while going for human protein (later you can cleave-but remember after cleavage it may immediately behave as different protein, I mean it aggregate unfortunately). Choose a E.coli which supply tRNAs for rarely used codons which needed for human proteins (Rosetta DE3 strains). Some says periplasmic expression may yield a fruitful amount, but I have no experience. So start with different pET/pGEXvectors and see where you are lucky enough to get your protein.

Good luck.

Dear Ananda,

I have already considered these issues and there is some solution in my system but i just want to know whether I can buy a expression system backbone for my protein or not. If i can buy it from some vendor, its great otherwise how can i make a new vector which i want to patent for my protein expression. Any suggestion!

You should consult a patent lawyer/scientist :) I am sorry I cant help you.

You can certainly buy it, but if it is covered by patents, you will not be able to commercialize it without entering into a licensing agreement. You'll need a tech transfer or patent lawyer for this.

To make your own plasmid you'll need replication origin, polyA signal, promoter sequence, antibiotic gene for selection. You might be able to use BioBricks (http://biobricks.org/) for open licensed parts to build your plasmid. This may reduce your ability to patent and monetize your product downstream, though.

If expression in bacterial proves difficult, you might try baculovirus expression in insect cells. Easily scalable, eukaryotic system that more easily expresses mammalian proteins with post-translational modifications - important for function in some proteins.

And, most importantly, talk to a lawyer before trying to start something commercial. You don't want to infringe someone's patent before you have a product.

u would want to look into the gateway system of invitrogen

Dear Matthew,

I am really very much thankful to you for your detailed and comparative reply. In my case, I want to use an expression vector and competent cells for some supplier like novagen or it may be some body other like NEB/ invitrogen etc. Here, i will make some modification in the vector backbone itself to express my protein of interest. Eventually, I want to take an IPR for the final product and the method used. I dont want to commercialize the vector. So, when i would like to transfer the technology to third party, what things are there that I must consider.

One more thing which i want to ask is.... What about the promoters? can i select any promoter from literature and it is possible that i can use this promoter in some other vector backbone. Or it is so that people have already IPRs on these promoters which prevent the use of these promoters by somebody else.

I am really very thankful to you and Research gate for this kind support and help.

Regards

-Yogendra

Dear Jason,

Thank you very much for your reply. I will try to see BioBricks, if it can help in understanding base structure.

Regards

-Yogendra

Dear Yogendra,

Did you get the solution to your problem. I was also looking for similar kind of problem. Please share your views.