Trying to expression a plant gene for amino acid synthesis in E. coli, but couldn't detect the functioning of this gene in E. coli and couldn't see a band in protein gel. What is the likelihood that this is a codon usage problem? What codon set is suitable fit for E. coli gene expression?
Hi Nanfei,
You need to see every step of your cloning and expression of plant gene in E.coli...as there can be several reasons why some protein may not express in E. coli..
1) Check the sequence of gene in the expression vector, for any mutations
2) Check for the codon usage of the E. coli and compare it to the gene sequence you are trying to over express.
3) You can try to express the protein in alternative E. coli hosts which are suitable for expression of Eukaryotic genes
4) See previous reports on this protein...it should not be problematic to E. coli
5) In such case you may also try invitro expression of protein
see whether you have tried to check these steps one by one......
best regards
Ajay Saini is true. I want to ask some questions. dose your expression vector have RBS? have you checked your interest gene RNA level in bacterial host?
Dear Colleague,
Simply check your gene sequence by getting sequencing. If there is no mutation then go for different IPTG concentration for expression.
- Badges
- Science topic
- Similar topics
- Biological Science
- Genetics
- Gene Expression