Hello to everyone. I have a question about the RNA extraction by using Trizol.
Almost everytime that I do this protocol, I obtain a very bad 260/230. This happens even if I have a good RNA concentration and a good 280/230.
Is that normal? Which could be the reason of that? Is there anything that I can do to improve it?
Thank you very much in advance!
You may try to to a RNA precipitation (with ethanol and Sodium acetate) procedure after the isolation with trizol, it may help improving RNA quality. It has helped me sometimes.
Hi Carla,
There are a few things you can try:
Make sure to shake vigorously after addition of chloroform (minimum 15 seconds if not more), this is vital to allow sufficient mixing and complete dissociation of nucleoprotein complexes allowing the proteins to be extracted to the organic phase.
Secondly go for quality over quantity when transferring the RNA supernatant, focus on retrieving RNA that is free from DNA and protein carryover.
These considerations may help you improve results
Hi Carla,
Whit Trizol is kid of hard to not get any DNA contamination in your RNA, and this could affect your measures.
We have been using is a DNAse kit to get rid of it, and it usually gives you a better yield, you can find it here http://www.thermofisher.com/order/catalog/product/AM1907
Before you try this, you could try making a PCR targeting DNA and check if it gives you an amplification.
Success in your research!
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