Hello, I am currently isolating nuclei from adipose tissue and I am interested in obtaining high-quality RNA for further assays. What extraction methods do you recommend?
I´m currently using the TRIzol protocol, without modifications but in addition to obtaining very little amount of RNA, the 280/260 and 280/230 values are very low, any suggestions?
For non-adipose tissue (i've worked predominantly with liver, lung, and kidney) I've always had great success with Applied Biosystems' MagMax RNA purification kit. I've used it for 100% manual purification and also for automated purification using the Kingfisher. And yes, RNA extraction from adipose tissue can be quite challenging, but try RNeasy lipid tissue mini kit by Qiagen (which i've also used with great success).
i agree with Emilee Bargoma RNA from tissue can be tough. are you getting good nuclear isolation? if so then use a lysis buffer with sodium dexoycholate (RIPA) to help punch holds in the membrane and lyse on ice for up to 30 minutes then perform RNA isolation. if you are not using a column then be very conservative when you are transferring phases and err on the side of less but clean RNA rather than trying to get every single ul of aqueous phase
- Badges
- Science topic