Hi,

I work with both HEK293f and HEK293T cell lines and I have no issues with RNA extraction in the respective cell lines. Try to use RNA mini kit from Invitrogen catalog #12183025.

For lysis buffer I use Trizol (600uL) and chloroform (200uL). I can send you the protocol if needed.

Generally speaking, seeding the cells in 6 well plate is enough for your RNA. And I prefer to harvest the cells when they reach 80-90% confluency.

Good luck in your experiment(s).

Hey Afi, I work with HEK293 cell line as well, the RNA extraction protocol I use is for TriZol reagent, maybe try this protocol (1ml TriZol reagent, Chloroform, Isopropanol and 75% total ethanol are the only reagents needed for this protocol). I get a high yield on the nano-drop after quantifying it. If you want I can also send through the protocol or you can find it online, if you just search Trizol RNA extraction protocol.

I aim to extract on the day when the cells are around 80-90% confluent for optimal yield in a 10cm plate.

With regards to the 260/230 ratio, make sure your area is sterile and you use RNAse away. To prevent any contamination.

I hope this helps.

All the best!