Need help in codon optimization... - (Dec/24/2014 )

I want to chemically synthesize a gfp gene for expressing in rice.I am using the IDT Codon Optimization program and found that not all the codons are 100% optimized.Some of them are the 2nd or 3rd in frequency of usage.Also the stop codon in the gene optimized is the least used in rice.I dont think all the codons can be the ones that are the most used in rice as there is need of translational pauses to get optimal protein expression.But how do I decide which codons are to be used in what positions to get maximum efficiency.Do I just use the automatic codon optimized gene given by IDT or further optimize it manually?

So should the stop codon used be the one which is the most frequently used in rice for proper mRNA termination?

Please help me as I am new to this and want tips and suggestions for getting optimized codons in this gene.

When we codon optimize, we calculate the frequency of codon use in the original organism, then match that frequency when we choose new codons in the target organism. The idea is to replicate things like translational pauses. You certainly don't want to have every codon be 100% optimized for a target organism. For GFP, this likely matters little, since GFP is s very simple straightforward protein. If you are concerned about stops, then you can add additional stops at the end of your construct. Reading through stop codons is much more common than people realize, and a second one will actually stop things.

You should look up the articles and patents by Claes Gustafson from DNA 2.0.