Computational protocol: RNA Sequencing Analysis and Atrial Natriuretic Peptide Production in Patients with Dilated and Ischemic Cardiomyopathy

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Protocol publication

[…] The initial whole transcriptome paired-end reads obtained from sequencing were mapped against the latest version of the human genome (version GRchr37/hg19) using the Life Technologies mapping algorithm (http://www.lifetechnologies.com/), version 1.3. It was using the standard Bioscope parameters of version 1.3, in paired ends and whole transcriptome analysis. For both reads, forwards and revers, the seed was the first 25 nucleotides with a maximum of 2 mismatches allows, and describe the main statistical parameters of mapping step of analysis. The aligned records were reported in BAM/SAM format . Bad quality reads (Phred score <10) were eliminated using Picard Tools software, version 1.83 .Subsequently, isoforms and gene prediction were estimated using the cufflinks method and the expression levels were calculated using the htseq software, version 0.5.4p3 , this method eliminate the multimapped reads, only the unique reads are consider for gene expression estimation. Edge method, version 3.2.4, was applied for differential expression analysis between conditions . This method rely on different normalize process based in depth global samples, CG composition and length of genes. In the differential expression process, this method relies on a Poisson model to estimate the variance of the RNA-seq data for differential expressions. The data discussed in this publication have been deposited in NCBI's Gene Expression Omnibus and are accessible through GEO Series accession number GSE55296 (http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE55296). […]

Pipeline specifications

Software tools Picard, Cufflinks, HTSeq
Databases GEO
Application RNA-seq analysis
Organisms Homo sapiens
Diseases Heart Failure, Cardiomyopathies