Porcine gene 1.1 st array strips

Total RNA was isolated from oxyntic and PYL collected from each subject, according to Qiagen RNeasy Midi Kit protocol (Qiagen, Hilden, Germany). To reduce the viscosity of the lysate, specimens (50 to 100 mg) were homogenized directly in the buffer RTL containing guanidine thiocyanate. All the other procedures were in agreement with the manufacturer protocol. Purity and integrity evaluation was assessed just before analysis by Agilent Bioanalyzer 2100. Total RNA was hybridized on Affymetrix Porcine Gene 1.1 ST array strips. Hybridized arrays were scanned on a GeneAtlas imaging station (Affymetrix, Santa Clara, CA, USA). Performance quality tests of the arrays including the labelling, hybridization, scanning and background signals by a Robust Multichip Analysis were performed on the CEL files using Affymetrix Expression Console. The intensity records were log2-transformed. Transcript data have been submitted to the National Center for Biotechnology Information's Gene Expression Omnibus (NCBI GEO) with GEO accession number GSE57620.

Total RNA was isolated from the gastric tissue samples using the FastPure RNA kit (TaKaRa Bio Inc., Shiga, Japan). All other procedures were in agreement with the manufacturer’s protocol. The RNA purity and integrity were evaluated using Agilent Bioanalyzer 2100 (Agilent Technologies, Palo Alto, CA) just before the transcriptome analysis. The total RNA was hybridized on Affymetrix Porcine Gene 1.1 ST array strips. The hybridized arrays were scanned on a GeneAtlas imaging station (Affymetrix, Santa Clara, CA, USA). Performance quality tests of the arrays including the labelling, hybridization, scanning and background signals using Robust Multichip Analysis were carried out on the CEL files using the Affymetrix Expression Console.

For microarray analysis, 6 out 10 pigs per group were randomly selected. Total mRNA was isolated from 50 mg of frozen distal jejunum mucosa according to the Takara Fast Pure kit (Takara Bio, Japan) protocol. The purity and concentration of the total RNA were checked using the Nanodrop ND 1000 (Nanodrop Technologies, USA), while RNA integrity was assessed by Agilent Bioanalyzer 2100 (Agilent Technologies, USA). Total mRNA was hybridized on Affymetrix Porcine Gene 1.1 ST array strips. Hybridized arrays were scanned on a GeneAtlas imaging station (Affymetrix, Santa Clara, CA, USA). Performance quality tests of the arrays including the labelling, hybridization, scanning and background signals by a Robust Multichip Analysis were performed on the CEL files using the Affymetrix Expression Console. The Affymetrix Transcripts ID’s, in general, characterized each one by several exonic sequences, were associated with 13,494 human gene names, based on Sus scrofa Ensembl (release 83, www.ensembl.org).