I'm trying to express a recombinant protein in CHO cells. In my first transfection experiment I couldn’t detect any protein expression (transfection efficiency was very low) and I was wondering if there is a place in the genome known for having high expression levels to lead my insert there by homologous recombination.
Using a reporter gene is a good option but, do you know any sequence that works well and do it faster?
Thanks for helping
We use the flp/in system for something like this. You transfect in a selectable plasmid with the FRT site and a very weak promoter on the selectable marker. Resistant cells tend do have the plasmid integrated in a favorable location for expression making the weak promoter strong enough to generate resistance. Once these cells are made you can recombine your gene of interest in using the FRT site and a site-specific recombinase. If you use a strong promoter on your gene it is now in a favorable genomic location.
Thank you, Prescott.
Decreasing the promoter's strength is very interesting. What kind of promoters do you use?
hey, usually mouse geneticists use the Rosa26 locus. There seems to be something similar in hamsters:
https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/SAFC/Posters/1/identification-of-the-putative-rosa26-locus.pdf
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