title: GSM8599322: SP1380 covS mutant strain in THY broth bulk RNA-seq 2; Streptococcus pyogenes; RNA-Seq
library_construction_protocol: RNA was isolated using the RNeasy Plus Kit (Qiagen). Briefly, bacteria were grown to mid-logarithmic growth phase in THY and pelleted. RNA was isolated from the dry pellet as per the manufacturer's instructions with an additional mechanical lysis step using Lysing Matrix B tubes (MP Biomedicals) on the FastPrep-2 5G bead beating grinder and lysis system (MP Biomedicals). On-column DNA extraction was also performed with RNase-Free DNAse Set (Qiagen) during RNA extraction according to manufacturer's instructions. To ensure complete removal of DNA, the RNA was then further purified using the TURBO DNA-free kit (Thermo Fisher Scientific) according to the manufacturer's instructions. RNAseq analysis was performed at the Australian Genome Research Facility (Melbourne, VIC, Australia). cDNA libraries were prepared from total RNA using TruSeq Stranded Total RNA Library Prep with Ribo-Zero Plus rRNA Depletion kit (Illumina). Sequencing of the cDNA libraries was performed on the NovaSeq 6000 System (Illumina) on a 2x150 bp SP flowcell run generating an average of 20 million reads per sample.