GSE113472
RNA Sequencing of si-Contol treated M010817 and siSMAD7 treated M010817 human melanoma cell line
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis Methods:  mRNA profiles of si-Contol treated M010817 and  siSMAD7 treated M010817 human melanoma cell line were generated by deep sequencing, in triplicate, using Illumina HiSeq 2500. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays Conclusions: Our study represents the first detailed analysis of si-Contol treated M010817 and siSMAD7 treated M010817 human melanoma cell line transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions. Overall design: mRNA profiles of si-Contol treated M010817 and  siSMAD7 treated M010817 human melanoma cell line were generated by deep sequencing, in triplicate, using Illumina HiSeq 2500.