Computational protocol: Trans-Reactivation: A New Epigenetic Phenomenon Underlying Transcriptional Reactivation of Silenced Genes

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Protocol publication

[…] Total long RNAs were extracted from malpighian tubuli tissues using the miRNeasy Mini Kit, following an RNeasy MinElute Cleanup Kit (Qiagen) step for the enrichment of small RNAs. The 100bp paired-end sequencing was performed at IGA using a HiSeq2000TM machine. Long RNAs were mapped against the Drosophila genome (assembly BDGP Release 5) using TopHat (PUBMED 19289445) with default parameters. Mapped reads were associated with gene annotations (FlyBase 5.12), and transcript levels of annotated genes, expressed as Fragments per Kilobase of exon Per Million fragments mapped (FPKM), were estimated by CuffCompare (PUBMED 20436464). For small RNAs, sequencing adapters were trimmed from sequence reads with custom scripts. Resulting sequences were mapped against the Drosophila genome using Bowtie (PUBMED 19261174) allowing at most two substitutions within the first 20 base pairs. […]

Pipeline specifications

Software tools TopHat, Cufflinks, Bowtie
Databases FlyBase BDGP
Application RNA-seq analysis