GSM7916844: tbsP_e_noglu; Haloferax volcanii DS2; RNA-Seq
After 24 hours storage in -80C, RNA was extracted from cell pellets using Absolutely RNA Miniprep kit (Agilent Technologies, Santa Clara, CA).  For all samples, the absence of DNA contamination was determined on 200 ng samples of RNA in 25 cycles of PCR and total RNA was quantified using Agilent Bioanalyzer RNA Nano 6000 chip (Agilent Technologies, Santa Clara, CA). For 2019 samples,  ribosomal RNA was removed with Ribo-Zero rRNA Removal Kit (now discontinued, Illumina, San Diego, CA). For 2021 samples, extraction was followed by additional DNAse treatment with Turbo DNAse (Invitrogen), and ribosomal RNA was removed from total RNA with the PanArchaea riboPOOL kit according to the manufacturer protocol (siTOOLs Biotech, Germany) and checked for depletion using Agilent Bioanalyzer RNA Nano 6000 chip as described as described in Pastor et al. 2022. For 2019 samples, rRNA-depleted RNA was used to construct the sequencing libraries with KAPA Stranded RNA-seq Library preparation (Kapa Biosystems) following manufacturer protocol. Fragment size of final libraries was measured using Agilent Bioanalyzer DNA 1000 chip and then pooled and sequenced on Illumina HiSeq4000 at the Duke Sequencing and Genomic Technologies Core Facility at Duke University (Durham, NC). For 2021 samples, sequencing libraries were constructed with NEBNext UltraII Directional RNA Library Preparation Kit (New England BioLabs, #E7760). The fragment size of all libraries was measured using Agilent Bioanalyzer DNA 1000 chip and then pooled and sequenced on NovaSeq6000 at the Center for Genomic and Computational Biology at Duke University (Durham, NC).