LINCATRA: Two-cycle method to amplify RNA for transcriptome analysis from formalin-fixed paraffin-embedded tissue

Abstract

Whole transcriptome analysis (WTA) using RNA extracted from Formalin Fixed Paraffin Embedded (FFPE) tissue is an invaluable tool to understand the molecular pathology of disease. RNA extracted from FFPE tissue is either degraded and/or in very low quantities hampering gene expression analysis. Earlier studies described protocols applied for cellular RNA using poly-A primer-based linear amplification. The current study describes a method, LINCATRA (LINear amplifiCAtion of RNA for whole TRAnscriptome analysis). It employs random nonamer primer based method which can amplify short, fragmented RNA with high fidelity from as low as 5 ng to obtain enough material for WTA. The two-cycle method significantly amplified RNA at ∼1000 folds (p 2, p < 0.05) identified between lung cancer and asthma cohorts validating the method developed. This method is applicable in clinical molecular pathology field for both diagnostics and elucidation of disease mechanisms.

Keywords

• Linear amplification
• Formalin-fixed paraffin embedded
• RNA-Seq
• Degraded
• Transcriptomic analysis