What housekeeper to use with HEK cells? - (May/17/2013 )

Hello everybody,

I am working with HEK293 cells and tried to check my RNA integration by using usual housekeepers (EF-1a, GAPDH, SDHD) but apparently they are not able to bind to this template, although in silico analyses revealed that they should. I tried them with quantitative PCR as well as with conventional RT-PCR with the same results.

I would be really happy for any suggestions.

Thanks a lot,

Best
Stephanie

Hmmm, those genes are all expressed by 293s, there is something wrong with either your PCR or your RNA extraction.

Thanks, yes, that's what I thought. We checked the RNA (it was fine), transcription was done in parallel with other RNA samples that all worked well, and the house keepers gave good results in a qPCR using tissues that was run the same day...

Btw: are there possibly any differences between transfected vs. non-transfected HEK cells in expressing these genes?

Potentially, you should test several housekeepers to find the ones (you should be using 2-3) that are most stable.