Illumina MiSeq paired end sequencing; Transcriptome profiling of Herbaspirillum rubrisubalbicans M1 during sorghum root colonization
Transcriptome profiling of Herbaspirillum rubrisubalbicans M1 during sorghum root colonization
Herbaspirillum rubrisubalbicans M1 was grown at 30ºC and 120 rpm in   NFbHPN medium (Klassen et al., 1997). Seeds of susceptible cultivar Sorghum   bicolor cv. A07 were surface-sterilized, germinated on water-agar (0.8%) plates and   transferred to a hydroponic system composed of 30 mL of plant medium (Egener et   al., 1999 without carbon and nitrogen sources) and 10 g of polypropylene spheres in   100 mL glass tubes (Balsanelli et al., 2013). Each seedling was then inoculated with   108 CFU of H. rubrisubalbicans. In the control condition without a plant, the   hydroponic medium was supplemented with 2 mM of malate. Bacterial samples   were taken 3 days after inoculation in duplicate. Free-living bacteria of the samples   without plants (named “control”) were isolated from 250 mL of hydroponic medium   by centrifugation (10,000 g, 8 min, 4°C) and re-suspended in cold lysis buffer (Tris   50 mM pH 8.0, EDTA 10 mM, SDS 0.6%). Epiphytic bacteria were removed from   roots by vortexing 10 g of roots in RNAlater (Sigma) during 5 min, isolated by   centrifugation (10,000 g, 8 min, 4°C) and re-suspended in cold lysis buffer. For both  samples (each in duplicate), total RNA was purified using TRI® Reagent (Sigma)   followed by chloroform extraction as suggested by the manufactor. Samples were   treated with turbo DNase (Ambion) and the RNA quantified with a NanoDrop   spectrophotometer (THERMO Scientific) and Bioanalyzer 2100 (Agilent). The   ribosomal RNA was partially depleted from 2 μg of total RNA using the Ribo Zero   Bacteria kit (Epicentre). The indexed cDNA libraries were constructed from 0.2 μg  ribosomal-depleted RNA samples using the TruSeq Stranded RNA Sample   Preparation kit v3 (Illumina).
Experimental Factor: growth condition
Sorghum root epiphytic cells
LIBRARY_STRAND
not applicable