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Old 05-29-2015, 03:18 PM   #3
kmcarr
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Location: USA, Midwest

Join Date: May 2008
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Quote:
Originally Posted by cmbetts View Post
You have plenty of RNA, so your main issue is the low RIN scores. The key issue with low RIN RNA is that using common PolyA purification methods to isolate mRNA will lead to extreme 3' bias and other issues in your libraries. However, if you use an rRNA depletion method (Lots kits using either probe pull-down or RNaseH digestion are available) and skip any fragmentation steps in your library protocol, you should be fine.
I'd recommend checking out the literature for RNA-Seq from FFPE RNA and use their methods for guidance.
Our lab recently used the NuGen Ovation RNA-Seq System for Model Organisms for some degraded mouse RNA sample (not FFPE, just lower quality) and it worked very well at eliminating rRNA. Unfortumately I don't have a comparable library prepared using a standard kit to assess any biases in mRNA representation.
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