E-MTAB-4958 - Xanthomonas campestris pv. Campestris 8004 far-red light irradiation and XccBphP overexpression transcriptional analysis
Submitted on 1 March 2016, released on 3 December 2016, last updated on 23 January 2020
Xanthomonas campestris pv. campestris
RNA isolation and RNA-Seq: Wild-type Xcc or pXccBphP strains were cultured in far-red light or dark conditions up to logarithmic phase (0.7 to 0.8 OD600) at 28C in PYM broth. Total bacterial RNA was isolated using the MasterPureTM RNA Purification Kit (Epicentre, Illumina). Samples corresponding to two biological replicates of each condition were submitted to Genome Qubec for rRNA removal with Ribo-Zero (Illumina) and TruSeq RNA-seq library preparation. 50 bp single-end sequencing of the libraries was performed using an Illumina HiSeq 2000 platform (Genome Qubec). Removal of low-quality reads and Illumina adapters, and assessment of the quality of the reads was performed using Trimmomatic (Bolger et al, 2014) and FastQC (www.bioinformatics.babraham.ac.uk/ projects/fastqc/), respectively. Rads were aligned to the Xcc genome obtained from GenBank (accession number: NC_007086.1) using SAMtools and the BurrowsWheeler Alignment software (BWA) (Li et al, 2009). Alignments were visualized using the software Integrated Genome Viewer (IGV) (http://broadinstitute.org/igv). RNA-Seq differential expression analysis: Read counts corresponding to annotated ORFs were quantified with the software FeatureCounts (Liao et al, 2014) using the strand specific mode. Differential expression analysis was performed using the software DESeq (Anders & Huber, 2010). Genes displaying adjusted p-value < 0.05.
RNA-seq of coding RNA, genetic modification design, growth condition design
Xanthomonas campestris attenuates virulence by sensing light through a bacteriophytochrome photoreceptor. Bonomi HR, Toum L, Sycz G, Sieira R, Toscani AM, Gudesblat GE, Leskow FC, Goldbaum FA, Vojnov AA, Malamud F.