RNA extraction from flat worms - (Nov/27/2013 )
I am extracting RNA from flat worms (Gyrodactylus salaris) with Trizol-based method, but I do not get any RNA, or RNA amounts are very low (the control, fish tissue samples are extracted normally).
Alive worms were frozen in liquid nitrogen in a drop of water (40 microliters). The number of worms per sample is around 30, and they are quite small (0.1 miliimeters). From this fresh-frozen samples I get nothing, whereas I was able to extract RNA from the same number of RNA-later stored worms previously.
I tried the standard protocol; additionally homogenizing the frozen drop of water to powder with immediate adding of Trizol; letting the samples to sit longer in Trizol for better lyzation; elongating the RNA precipitation step, but nothing seems to work.
Does anybody have an experience with extracting from very small initial amounts of tissue or may suggest something?
I will appreciate your help!!
You could try flash freezing and pulverizing. Best would probably be freeziing in LN2 or dry ice/ethanol, followed by grinding with a bead beater in a small volume of trizoi. I would add the trizol directly to the frozen sample pellet before the grinding.
Thank you! Actually, that what I did initially, with no success however..