GSM5002060: Lid S1; Homo sapiens; RNA-Seq
Total RNA was isolated from ten to fifteen formalin-fixed and paraffin-embedded (FFPE) 4-µm-thick sections of all specimens using the Quick-RNA FFPE Kit (Zymo Research, USA). Following a DNAse I digestion using the Baseline-ZERO kit (Epicentre, USA), the RNA concentration was measured with the Qubit RNA HS Assay Kit on a Qubit Fluorometer (Life Technologies, USA). The RNA quality was determined with the RNA Pico Sensitivity Assay on a LabChip GXII Touch (PerkinElmer, USA). The fragment size of all RNA samples ranged between 120 and 150 bp. RNA sequencing was performed using massive analysis of cDNA ends (MACE), a 3' RNA sequencing method. The preparation of MACE libraries was carried out using 1 µg of total RNA, as previously described. The barcoded libraries were sequenced simultaneously on the NextSeq 500 (Illumina, USA) with 1× 75 bp. PCR bias was removed using an unique molecular identifier.