Hi,
My question will be at a very fundamental level about heterologous gene expression,
For any (heterologous) gene we insert between any promoter and the corresponding terminator sequence, is it possible to express it successfully at low or high levels in a host cell?
For heterologous expression, should I either clone any gene with its natural promoter and terminator sequences or insert it into the 'coding region' of a P+coding region+T sequence already existing/working in the host cell's genome?
Many thanks,
In general, you should rely on the host control elements (promoter group, operators, terminators) rather than to bring them from another organism. Also, it's advisable to adjust the coding sequence of your recombinant protein to the codon usage frequencies of the host.
However, if your recombinant sequence originates from an organism which is phylogenetically close to the host, there is a good chance that you can transfer it together with promoters/operators/terminators, and such an expression unit will still work.
Abdurrahman Aygül It means, Targeted knock-in vs Stable-transgenic expression.
Targeted knock-in - Difficult to perform, can use native elements, less prone to genomic silencing, mostly weaker expression than the TG.
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