subject status: patients diagnosed with gastric cancer patient id number: 14 tissue type: primary tumor
Extracted molecule
total RNA
Extraction protocol
Pairs of primary tumours and corresponding normal gastric mucosa were sampled from patients’ surgical specimens immediately after resection. The fresh specimens were collected into RNase free polypropylene tubes filled with RNAlater® stabilization solution (Qiagen, Valencia, CA, USA) and stored at −80 °C until RNA was extracted. Total tissue RNA was extracted using miRCURY RNA Isolation Kits (Exiqon, Denmark) according to the manufacturer’s instructions. RNA concentrations were measured with NanoDrop (Thermo Fisher Scientific, Waltham, MA, USA) and overall sample quality was determined by RNA integrity number (RIN) using Agilent Bioanalyser (Agilent Technologies, Palo Alto, CA, USA).
Label
Cy3, Cy5
Label protocol
Total RNA (750 ng) isolated from tumour and healthy mucosa samples was labelled with Hy3™ and Hy5™ fluorescent labels, respectively, using the miRCURY LNA™ microRNA Hi-Power Labelling Kit, Hy3™/Hy5™ (Exiqon, Denmark) following manufacturer's instructions.
Hybridization protocol
The labelled RNA samples were hybridized to the miRCURY LNA™ microRNA Array 7th Gen (Exiqon, Denmark) containing capture probes targeting all microRNAs of human origin registered in the miRBASE 19.0. The hybridization was performed using a Tecan HS4800™ hybridization station (Tecan, Austria) according to the manufacturer’s protocol.
Scan protocol
After hybridization, the microarray slides were scanned using the Agilent G2565BA Microarray Scanner System (Agilent Technologies, USA)
Description
14
Data processing
Microarrays were analysed with the ImaGene® 9 software (BioDiscovery, CA, USA). The obtained signals were background corrected and data normalization was performed with the Lowess (Locally Weighted Scatterplot Smoothing) algorithm. MicroRNAs with intensities above threshold in less than 20% of the samples were removed from the final dataset used for the expression analysis. The average_FC_data.txt contains the following data columns; logFC – log FoldChange of individual microRNAs of cancer vs healthy mucosa AvgExpr – average expression for a given microRNA AvgHy3 – average Hy3 signal well above background P.Value – raw P value adj.P.Val – adjusted P value
