Five micrograms of each cDNA was fragmented and biotin-labeled using the Encore Biotin Module and instructions (Nugen).
RNA samples were amplified by using Ovation PicoSL System V2 (NuGene Technologies, CA, USA) and ENCORe Biotin module (NuGene Technologies).
Products purified from single primer isothermal amplification (SPIA) were then fragmented and labeled with biotin using Encore Biotin Module (NuGEN).
Colonic mucosal RNA was isolated by Qiangen-Qiazol - miRNA Isolation Kit. cDNA amplification and labeling was performed with Ovation Pico WTA System V2 and Encore Biotin Module (NuGEN), respectively.
cDNA amplification (Ovation Pico WTA systems, NuGEN, San Carlos, CA), labelling and hybridization (Encore Biotin Module, NuGEN) were carried out by the microarray core facility at University of Pennsylvania.
After purification according to the manufacturer's protocol, 2.5 μg of single-stranded DNA was fragmented and labeled with biotin using the Encore Biotin Module (NuGEN).
The obtained SPIA-amplified cDNA was purified using Agencourt RNA clean XP Purification Beads and fragmented (5 ng) and labeled with biotin using the Encore Biotin Module (NuGEN, San Carlos, California).
For microarray analysis, cDNA was prepared using the Ovation Pico WTA System V2 and labeled using the Encore Biotin Module (NuGEN, San Carlos, CA).
Total RNA from freshly sorted HMECs from donors M3, M6, M8, M9, M10 and M12 was amplified using the Ovation Pico WTA System V2 in combination with the Encore Biotin Module (Nugen).
Resultant cDNA probes were labeled with the Encore Biotin Module (NuGen Technologies, Inc). and hybridized to the Affymetrix GeneChip Mouse Gene 1.0 ST Array according to the manufacturer's instructions.
RNA was amplified with Nugen Ovation PicoSL WTA system (Cat nr. 3310-48), labeled with the Encore BiotinIL Module (Cat nr. 4210-48) and hybridized to Illumina MouseRef8 bead-chip microarrays.
