Sureprint g3 human gene expression 8x60k v2 microarray kit

Three sets of biological replicates of untreated HeLa and DIA-treated HeLa were prepared. HeLa cells were seeded in 6 well plates at a density of 2.4 × 10⁵ cells/well overnight. After 24 h, 15 μg/mL of DIA was added into the designated wells and was left to incubate for 48 h in a humidified 37°C CO₂ incubator. After the incubation time, the cells were harvested and RNA was extracted using the QiagenRneasy Mini Kit (Qiagen, Germany). The quality of the RNA extracted was measured using the 2100 Bioanalyzer using a RNA Pico chip (Agilent, USA). In order to proceed to microarray, the RIN (RNA Integrity Number) should be more than eight. After all of the samples have passed the minimum requirement for microarray analysis the samples were then used for microarray. All of the samples were subjected to the SurePrint G3 Human Gene Expression 8x60K v2 microarray kit (Agilent Technologies, USA) according to manufacturer protocol, and scanned with Agilent DNA microarray scanner. The results from the microarray study has already been uploaded on the Gene Expression Omnibus with the accession number GSE72974.

To identify genes regulated by LINC-PINT, LINC-PINT-overexpressing HNE1 cells and control group cells were selected for microarray analysis. In Brief, total cellular RNA was extracted using the RNAiso Reagent (Takara, Japan), and qualified RNA was then purified by mirVana miRNA Isolation Kit (Ambion, Austin, TX, USA). After purification, cDNA was amplified using a poly dT-T7 promoter primer and covalently coupled with Cyanin3 (Cy3). Labeled cDNA samples were then subjected to SurePrint G3 Human Gene Expression 8x60K v2 Microarray Kit (Agilent). For downstream analysis, gene array data were analyzed and filtered using the GeneSpring software V13 (Agilent) according to the manufacturer’s instructions. Relative to control, signal intensity >onefold change and P < 0.05 were considered as differentially expressed genes, which were induced by LINC-PINT overexpression. For functional gene enrichment, GO analysis was performed and the P value was calculated by Bonferroni corrected Fisher’s exact tests.