GSM4787257: L. interrogans mutant Man1474 with an inactivated LIMLP_10155 (perRA) gene replicate #1; Leptospira interrogans serovar Manilae; RNA-Seq
From 0.5 μg of total RNA, we depleted ribosomal RNAs using the Ribo-Zero rRNA Removal Kit (Bacteria) from Illumina. On these rRNA-depleted RNAs, the sequencing libraries were constructed using the TruSeq Stranded mRNA Sample preparation kit following the manufacturer's instructions (Illumina). The directional libraries were controlled on Bioanalyzer DNA1000 Chips (Agilent Technologies) and concentrations measured with the Qubit® dsDNA HS Assay Kit (ThermoFisher). Sequences of 65 bases were generated on the Illumina Hiseq 2500 sequencer. Harvested cells were resuspended in 1 ml TRIzol (ThermoFisher Scientific) and stored at -80°C. Nucleic Acids were extracted with chloroform and precipitated with isopropanol as described earlier (zavala-Alvarado et al., 2020). Contaminating genomic DNA was removed by DNAse treatment using the RNAse-free Turbo DNA-freeTM turbo kit (ThermoFisher Scientific) as described by the manufacturer. The integrity of RNAs (RIN > 8.0) was verified by the Agilent Bioanalyzer RNA NanoChips (Agilent technologies, Wilmington, DE).