Formalin-Fixed, Paraffin-Embedded (FFPE) material is very heterogeneous and highly variable in terms of quality, degree of cross-linking, and accessibility of the mRNA. Therefore, processing degraded or FFPE RNA with QuantSeq-Pool may lead to uneven read distribution between samples in sequencing experiments.
For best practice, the QuantSeq FWD 3' mRNA-Seq V2 Library Prep Kit for Illumina with UDIs (Cat. No. 015, 113 – 115, 129 - 131191 - 196) is recommended for processing FFPE samples, as it allows quality control and quantification of individual libraries.
This offers the possibility to re-adjust individual samples for equimolar lane pooling and balanced read distribution during sequencing.