Genome-Wide miRNA Profiling in FFPE Tissue

In our discovery phase we conducted genome-wide miRNA profiling using Taqman MicroRNA Arrays, also known as Taqman Low Density Array (TLDA) ‘Pool A’ Card version 3.0, due to their established use for miRNA expression analysis in FFPE tissue [28 (link),29 (link)]. This 384-microfluidic array was designed to perform quantitative reverse transcriptase (qRT-PCR) reactions simultaneously (Applied Biosystems, Austin, TX, USA) on 378 mature miRNAs (and 6 endogenous controls) that tend to be functionally defined. Using 20 nanograms (ng) of total RNA as input, cDNA was synthesized with highly multiplexed Megaplex RT primers, pre-amplified with Megaplex PreAmp Primers, mixed with TaqMan Universal PCR Master Mix (Applied Biosystems), and loaded onto TLDA cards for expression analysis.

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Permuth-Wey J., Chen Y.A., Fisher K., McCarthy S., Qu X., Lloyd M.C., Kasprzak A., Fournier M., Williams V.L., Ghia K.M., Yoder S.J., Hall L., Georgeades C., Olaoye F., Husain K., Springett G.M., Chen D.T., Yeatman T., Centeno B.A., Klapman J., Coppola D, & Malafa M. (2015). A Genome-Wide Investigation of MicroRNA Expression Identifies Biologically-Meaningful MicroRNAs That Distinguish between High-Risk and Low-Risk Intraductal Papillary Mucinous Neoplasms of the Pancreas. PLoS ONE, 10(1), e0116869.

Publication 2015

Megaplex RT primers Megaplex PreAmp Primers TaqMan Universal PCR Master Mix

Austin Cdna Mirnas Primers Reverse transcriptase Tissue

Corresponding Organization : Moffitt Cancer Center

Other organizations : Spartanburg Regional Healthcare System

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Variable analysis

independent variables

Taqman MicroRNA Arrays, also known as Taqman Low Density Array (TLDA) 'Pool A' Card version 3.0

dependent variables

MiRNA expression analysis in FFPE tissue

control variables

20 nanograms (ng) of total RNA as input
CDNA synthesized with highly multiplexed Megaplex RT primers
Pre-amplified with Megaplex PreAmp Primers
Mixed with TaqMan Universal PCR Master Mix (Applied Biosystems)
Loaded onto TLDA cards for expression analysis

controls

6 endogenous controls

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