Agilent s sure hyb hybridization chambers

To screening the potential targets of miR‐192 and ‐215, gene microarrays were carried out on the Agilent Whole Genome Oligo Microarrays (4x44K, Agilent, Santa Clara, CA, USA) in GC cells. All the procedures were referred as the manufacture protocols. Briefly, microarrays performance and analysis were performed on two groups of cell lines: BGC823 cells with loss‐function of miR‐192 and ‐215, and HFE145 cells with gain‐function of miR‐192 and ‐215. Lyse cells directly in a culture dish by adding 1 mL of TRLzol Reagent (Invitrogen, Carlsbad, California, USA) to a 3.5 cm diameter dish, and passing the cell lysate several times through a pipette. The amount of TRIzol Reagent added is based on the area of the culture dish (1 mL per 10 cm²). The quality and quantity of the RNA samples were checked, using an ND‐1000 Spectrophotometer (NanoDrop Technologies, Wilmington, Delaware, USA). RNA Integrity and cDNA contamination were tested by Denaturing Agarose Gel Electrophoresis. Agilent Quick Amp Labeling Kit and Agilent's Sure Hyb Hybridization Chambers (Agilent, Santa Clara, CA, USA) were used for sample labeling and hybridization, respectively. After scanning, data were extracted and further analysis was performed, using Agilent GeneSpring GX 11.5.1 software.
Upregulated and downregulated genes were identified by the threshold with at least twofold between the two groups as significance.

To know the targets of miR-192/215, we performed gene microarrays, which were carried out by the Agilent Whole Genome Oligo Microarrays (4×44K, Agilent, Santa Clara, CA, USA). Different expression genes were identified between two groups of cells, BGC823 cells with inhibition of miR-192/215, and HFE145 cells with mimics of miR-192/215. Upregulated or downregulated at least two-fold genes were counted as significance. Briefly, TRIzol Reagent (Invitrogen, Carlsbad, California, USA) lyses cells, and RNA Integrity was tested by Agarose Gel Electrophoresis. Agilent Quick Amp Labeling Kit and Agilent’s SureHyb Hybridization Chambers (Agilent, Santa Clara, CA, USA) were applied for the sample labeling and hybridization as per the manufacture protocols. Data were extracted and further analysis was performed by Agilent GeneSpring GX 11.5.1 software.