Computational protocol: Melatonin suppresses hepatocellular carcinoma progression via lncRNA-CPS1-IT-mediated HIF-1α inactivation

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Protocol publication

[…] After removing adapter sequences from the raw data, the individual library was converted to FASTQ format. Sequence reads were aligned to the human genome (hg19) with TopHat2 [], and the resulting alignment data were reconstructed with Cufflinks [] and Scripture []. The RefSeq database (Build 37.3) and GENCODE v19 database were used as annotation references for mRNA and lncRNA analyses, respectively. Protein coding potential of transcripts was further analyzed by Coding Potential Calculator [], Pfam-scan [], phylogenetic codon substitution frequency [], and Coding-Non-Coding-Index []. Transcript assembly, abundance estimates, and differential expression analyses were conducted by using Cufflinks2 and Cuffdiff2 [, ]. Biological sample gene variance was not determined because differential gene expression comparison was run without biological replicates. Gene or exon expression levels were normalized to the number of reads per kilobase per million mapped reads []. An adjusted p value < 0.05 (Student’s t-test) was used as the cut-off for determining differentially expressed genes. […]

Pipeline specifications

Software tools TopHat, Cufflinks, Scripture, CPC, CNCI
Databases Pfam GENCODE
Applications RNA-seq analysis, Transcription analysis
Diseases Carcinoma, Hepatocellular, Neoplasms
Chemicals Melatonin