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RNA-Seq analysis of melanoma cells, B16F10, in the presence of hypochlorous acid (HOCl)

ABSTRACT: RNA-Seq analysis of melanoma cells, B16F10, in the presence of hypochlorous acid (HOCl)

PROVIDER: PRJNA688512 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR13324777_1.fastq.gz	Fastqsanger.gz
	SRR13324777_2.fastq.gz	Fastqsanger.gz
	SRR13324778_1.fastq.gz	Fastqsanger.gz
	SRR13324778_2.fastq.gz	Fastqsanger.gz
	SRR13324779_1.fastq.gz	Fastqsanger.gz

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RNA-Seq analysis of melanoma cells, B16F10, in the presence of hypochlorous acid (HOCl)

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Project description:Hypochlorous acid (HOCl) is a potent oxidant that is produced endogenously in mammalian tissue by phagocytes. Exogenous exposure to HOCl also can occur following inhalation of chlorine gas. HOCl has been implicated as a source of oxidative stress associated atherosclerosis and other diseases. The purpose of this study was to identify dose-dependent transitions in cellular response to hypochlorous acid (HOCl), with a focus on understanding how various cellular defense and stress dose-responses overlap. Experiment Overall Design: RAW 264.7 cells were grown to 80-90% confluency in DMEM supplemented with supplemented with 10% fetal bovine serum (FBS), 10 mM HEPES, 100 U penicillin/ml, and 100 Âµg streptomycin/ml. Cells were treated with 0, 0.14, 0.35, 0.7, 1.4, 2.1, 2.8 or 3.5 mM HOCl in medium for 6 hr. Total RNA was isolated from cells with TRIzol according to manufacturer's instructions and then subjected to cleanup using RNase-Free DNase Set and RNeasy Mini kit. The resultant DNA-free RNA was diluted in RNase-free H2O and quantified by Nanodrop at 260 nm. The quality of RNA samples was confirmed using RNA Nano Chips with Agilent 2100 Bioanalyzer. RNA samples were stored at â70 oC until use. Experiment Overall Design: From 5 micrograms of total RNA, cDNA was synthesized using a one-cycle cDNA synthesis kit (Affymetrix Corp., Santa Clara, CA). cDNA was transcribed to cRNA which was then biotin-labeled using GeneChip IVT labeling kit. Fifteen micrograms of labeled cRNA were then hybridized to an Affymetrix Mouse Genome 430 2.0 Array at 45Â°C for 16 hr. Biological cRNA replicates (n = 3) were each hybridized to an individual array. After being washed using the GeneChip Fluidics Station 450, arrays were scanned using a GeneChip 3000 scanner and intensity values were extracted from the CEL file using Array Assist software (Stratagene, La Jolla, CA). Experiment Overall Design:

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