Computational protocol: Transcriptomic Analysis Reveals Adaptive Responses of an Enterobacteriaceae Strain LSJC7 to Arsenic Exposure

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Protocol publication

[…] Clean data were obtained using in-house perl scripts by removing reads containing adapter, reads containing ploy-N, and low quality reads from raw data of FASTQ format. RNA-Seq reads (clean data) were aligned to the LSJC7 reference genome. Both building index of reference genome and aligning clean reads to reference genome were used in Bowtie2-2.0.6 (). HTSeq v0.5.4p3 was used to count the reads numbers mapped to each gene. And then Reads Per Kilobase of exon model per Million mapped reads (RPKM) of each gene was calculated based on the length of the gene and reads count was mapped to this gene ().The read counts of each sequenced library were adjusted by edgeR program package () through one scaling normalized factor. Differential expression analysis of two conditions was performed using the DEGSeq R package (1.12.0; ). The P-values were adjusted using the method. Corrected P-value of 0.005 and log2 (Fold change) of 1 were set as the threshold for significantly differential expression.Gene Ontology (GO) enrichment analysis of differentially expressed genes was implemented by the GOseq R package, in which gene length bias was corrected (). GO terms with corrected P-value less than 0.05 were considered significantly enriched by differentially expressed genes. […]

Pipeline specifications

Software tools Bowtie2, HTSeq, edgeR, DEGseq, GOseq
Application RNA-seq analysis
Chemicals Arsenic, Nitric Oxide, Oxygen