Dasl ht assay

Total RNA was isolated, labeled and hybridized using the Whole-Genome protocol from DASL HT Assay (Illumina, San Diego, CA, United States) and Ovation^® FFPE WTA-Systems (NuGEN, San Carlos, CA, United States) to an Illumina humanRef-12 beadarray (Illumina, San Diego, CA, United States) according to the manufacturer’s protocol. Raw bead count data was analyzed using the R/Bioconductor package beadarray (Dunning et al., 2007 (link)) followed by quantile normalization and log2 transformation. Data is accessible on Gene Expression Omnibus (GEO) as GSE101448 and GSE101462. Public microarray data from GEO were downloaded in pre-processed form, i.e., normalized, log2 transformed and filtered on probe level, according to the original reference (see Table 1). Since E-MEXP-1121 is a follow-up study of E-MEXP-950, we combined the two data sets from ArrayExpress. HGU133a and HGU133b chips from both studies were normalized by robust multiarray averaging (RMA) and subsequently combined, neglecting the lower inter-quartile range (IQR) if probes were present on both chips. For all data sets platform probe IDs were matched to unique EntrezIDs. In the case of multiple probes matched the same EntrezID, we chose the probe with the largest IQR.

This data set consists of six normal colon samples, six microvesicular hyperplastic polyps (MVHPs) and six sessile serrated adenomas/polyps (SSA/Ps) [32 (link)]. The total RNA was converted to cDNA and modified using the Illumina DASL-HT assay and hybridized to the Illumina HumanHT-12 WG-DASL V4.0 R2 expression beadchip. The biopsies were classified by seven gastrointestinal pathologists who reviewed 109 serrated polyps and identified 60 polyps with consensus. The log₂-scale of the expression measurements provided under the gene expression omnibus (GEO) accession number GSE43841 was used. Only MVHP and SSA/P samples were considered for the analyses. Illumina probe identifiers were mapped to gene symbol identifiers using the Bioconductor annotation package illuminaHumanWGDASLv4.db. Whenever multiple probes were mapped to the same gene, the probe with the largest t-statistic between MVHP and SSA/P was selected.