Usually RNA is extracted from fresh frozen tissue. is it frozen?

http://www.biomedcentral.com/1471-2199/8/118

Yes, it was kept in -80 for 2 and a half years. I am suspecting the time length of the leaf storage could be reason for no RNA

.

Did you keep the sample deep frozen during the preparation for RNA extraction? For instance, from the freezer to a mortar chilled with liquid N2 and then grind the leaf to a powder.

The leaf was already grinded and put in the tube for long. I took the sample out from -80 and added trizol immediately to remove any RNase activity.

Hi Faraz,

Do you need to extract total RNA or do you need to amplify a specific gene of partial gene region?

Regards,

Chris

Sometimes with Trizol extraction you will have no visible pellet - but the RNA is indeed there.  After brief drying of the pellet, use a small amount of nuclease-free water to resolubilize (10 to 30 uL).

page 19 and 20 of the PDF at the following link is a good approach...

http://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=1&ved=0CB4QFjAA&url=http%3A%2F%2Fwww.springer.com%2Fcda%2Fcontent%2Fdocument%2Fcda_downloaddocument%2F9781588294302-c2.pdf%3FSGWID%3D0-0-45-383626-p173729440&ei=lAvyVIfdDI7msASj7IDgCQ&usg=AFQjCNHnJuMd7acG89VzX2nlk1ijKrwfjA

Hi Chris,

I need to extract the total RNA for qPCR analysis.

Thanks Jack. It was helpful. 

Good~!  Sometimes not being able to see the pellet makes one doubt anything is there.  You just have to work through that doubt to the end of the protocol each time is all.

Hi Jack. That's what i did and it worked out well. Only difference that the amount of conc. i got was a bit low but i can work with it. Thanks :)