Hi,
For E. coli, as others already pointed out - optimizing codon usage of a gene 'does not usually hurt'. I can attest to the fact that it can boost expression quite a bit - at least in the modest number of cases where I have had a chance to compare the expression levels under the same conditions. However, it isn't easy to separate the effect of codon optimization from that of the mRNA structure changes. Most synthetic DNA companies boast their own ways of designing 'optimized DNA' - some do Monte Carlo codon substitution, others try to predict (and eliminate) local stem-loop structures by sliding a sequence->structure window of various size along the mRNA strand. I am sure there are other ways of doing this as well. The end result is a reshuffled DNA sequence. There also are companies who claim to be able to design optimized mRNA for specific expression needs, since it is known that not all mRNA structure is bad, and that translational pausing is important for the proper folding of certain polypeptides. For yeast and chloroplast expression codon use matching to that of the host organism(elle) is essential, based on the growing number of reports and on personal experience. For mammalian cell expression the situation is not as clear. In E.coli it is reasonably easy to control expression levels by changing growth and induction parameters - so I wouldn't worry about making too much gene product - since you can almost always dial the expression down (I wish it was as easy to dial it up!). Happy expressionizing. Artem _____ From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Looney, Andrea Lynn (Andrea Hevrdeys) Sent: Friday, February 01, 2008 10:29 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Codon Optimized Expression Dear All, I am getting very little (not zero) expression of my protein. I am curious to know if it is worthwhile to codon optimize my gene, which is ~1200bp, for E.coli. Can you have "too much of a good thing"? Can codon optimizing overwhelm the cell or in some way cause death or lowered expression? Thank you all, Andrea L. Hevrdeys
