Transcriptomic Analysis of ES Biopsies

Transcriptomic analysis was carried out using the SURMetaGIT protocol^{10 (link)}. In brief, following RNA extraction (RNeasy Plus kit-Qiagen, Germantown, MD), ES biopsies with an RNA concentration ≥100 ng/µL, and RNA integrity number (RIN) ≥7, were submitted for global expression analysis using the Human GeneChip 1.0 ST Array (Affymetrix, Inc., Santa Clara, CA). Significance of microarrays and rank product methods were analysed to select differentially expressed genes, using p < 0.05 (corrected for false discovery rate). Target analysis was also performed by real time quantitative polymerase chain reaction (RT-qPCR) to validate certain genes of interest which are potentially involved in carcinogenesis. We did this by using TaqMan gene expression assays (Thermo Fisher Scientific, Waltam, MA). β-actin was used as a reference gene.