Computational protocol: Comparative analysis of transcriptomes in aerial stems and roots of Ephedra sinica based on high-throughput mRNA sequencing

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Protocol publication

[…] The RNA-Seq reads in fastq format were assembled using the Rnnotator program and contig sequences were output in fasta format. Searches by blastx query with an E-value cutoff of 1E-6, GO mapping, and annotation by EC and IPR numbers were performed for Es_S, Es_R, and combined Es_S and Es_R (Es_SR) contigs continuously using the Blast2GO program , , . The method for quantitation of gene expression levels in the aerial stems and roots is summarized in . In this expression analysis, mapping of short sequence reads in fastq format of Es_S and Es_R to Es_SR contigs was performed using TopHat . The gene expression levels in the Es_S and Es_R transcriptomes were quantified by using Cufflinks software, and the abundances of expressed genes were calculated as expected fragments per kilobase of transcript per million fragments mapped (FPKM) . The differential gene expression levels of the Es_SR combined transcriptomes in Es_S and Es_R were quantified using Cuffdiff in the Cufflinks program . The significance of the abundance of an expressed gene was determined by the false discovery rate < 5% (q value < 0.05). […]

Pipeline specifications

Software tools Rnnotator, BLASTX, Blast2GO, TopHat, Cufflinks
Application RNA-seq analysis