mRNA Profiling of miR-17 family inhibition using TuD lentiviral vector in HepG2 and SK-Hep1 hepatocellular carcinoma cell lines [RNA-Seq]

To functionally characterize the role of miR-17 family in HCC, lentiviral vector-based miR inhibitor TuD was used to inhibit miR-17 family of microRNAs in HepG2 and SK-Hep1 HCC cell lines Overall design: Methods: HepG2 and SK-Hep1 HCC cell lines were acquired from American Type Culture Collection (ATCC) and miR-17 TuD or NC TuD expressing lines were generated. mRNA profiling of miR-17 TuD or NC TuD expressing samples was performed using Illumina NGS. Total RNA was extracted as per manufacturer’s instructions (RNeasy kit, Qiagen). RNA quality was assessed using BioAnalyzer (Agilent). mRNA expression profiles were determined using next-generation sequencing (NGS) on the Illumina HiSeq 2000 platform producing 50bp paired-end reads. Bowtie/TopHat suites were used to align the reads to mouse genome or transcriptome and RSEM were used to quantify gene abundances. Gene level counts were then normalized with the R/Bioconductor package limma using the voom/variance stabilization method.

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Huang X, Magnus J, Kaimal V, Karmali P, Li J, Walls M, Prudente R, Sung E, Sorourian M, Lee R, Davis S, Yang X, Estrella H, Lee EC, Chau BN, Pavlicek A, Zabludoff S