HELP- Can I save my RNA now? - RNAse free pipette tips (Jun/27/2008 )
I am new to PCR, and I have some valuable samples to take out the RNA from (cells). I have scraped them in TRIZOL and now they are sitting at -80. The thing is I forgot to use RNAse free pipette tips while putting the TRIZOL in. I used just simple autoclaved tips. Can my samples be saved now? Does nyone have experience with this? How much RNA would have degraded? Please help, these samples were very valuable.
With Trizol you don't even need autoclaved tips. You are fine.
Why wouldn't he need RNAse free tips with Trizol?
I still think you're fine though
I still think you're fine though
Trizol contains guanidine thiocyanate, which I believe should inactivate RNases as well. So you're good to go
yes it is right, dont be worry about it,
but you'd better use these -80 fast as Rnase is stable and could be refold in lower concentration of guanidine thiocyanate.
cheers,
akhshik.
maybe your normal-autoclave pipette tips do not contaminated with RNase... then your RNA will be fine.
my friend once try using her normal-autoclave tips, tubes, etc etc... and do not wipe her things with RNase Away.
She got good RNA with no degradation or DNA contamination.
We treat things with DEPC or use RNase free stuffs to prevent any Rnase contamination "JUST-IN-CASE" there are any contamination.
the possibility of RNase contamination is there, but not definately. sometimes, if you are aseptic enough and you have a good technique, you won't get your RNA degrade. But, when we deal with RNA, we really don't want to take any risk. therefore we always use RNase free stuffs.
Hey! You're are definetely fine!
Did my whole PhD doing RNA extractions for downstream Northern blot and qPCR using only autoclaved material, my lab didn't have the cash to buy fancy RNase-free filtered tips, etc, RNase-awya, or anything like that, and we very rarely had a problem with degradation of RNA.
Good luck!
As everyone has said - you have not problem at all. One logic to keep in mind regarding using RNase free stuff is to ask if your stuff - tips, tubes, homogenizer, cell suspension buffer, etc. - would have more RNases than the cell culture or tissue you are processing. Of course not. RNA is only in danger after it is purified and no longer protected by the extraction buffer. There is no need to waste time and money, and worry in your case, for "RNase-free" in the early steps of RNA extraction.