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RNA extraction from flat worms

RNA extraction worms small yield Trizol

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#1 Ksenia

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Posted 27 November 2013 - 01:46 AM

Dear all,

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!!

 



#2 phage434

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Posted 27 November 2013 - 06:50 AM

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.



#3 Ksenia

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Posted 28 November 2013 - 05:30 AM

Thank you! Actually, that what I did initially, with no success however..







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