GSM1161283: FS; Mycobacterium tuberculosis H37Rv; RNA-Seq
Total RNA was isolated from cultures in exponential and stationary phases. Basically, cultures were harvested by centrifugation and pellets were resuspended in Guanidinium Chloride buffer (6M guanidinium chloride, 0.1% Tween 80, 1mM, 2-ß-mercaptoethanol, 10mM EDTA) in a proportion of 1ml of buffer per 100 ml of culture. Cells were lysed mechanically in a FastPrepTM (Thermo Scientific) using 150-200 µm glass beads (Sigma Aldrich) by doing 4 lysis cycles at high speed (6.5 m/s) during 15 s each. Nucleic acids were purified with phenol-chloroform-isoamyl alcohol (25:24:1) and RNA was differentially precipitated with 0.4 vol of absolute ethanol (added drop by drop). Finally, RNA was washed 3 times with TrizolTM reagent (Invitrogen). RNA integrity was analyzed using a bioanalyzer (Agilent technologies) and quantified by spectrophotometer using the nanodrop ND-1000 (Thermo Scientific). Directional RNA-seq, Waldbauer J.R et al (2012)
GEO Accession
GSM1161283