title: HiSeq X Ten paired end sequencing; Comparison of the transcriptional profiles of wild-type and ΔAvin_01720 Azotobacter vinelandii strains cultured under aerobic nitrogen-fixing conditions
design: Comparison of the transcriptional profiles of wild-type and ΔAvin_01720 Azotobacter vinelandii strains cultured under aerobic nitrogen-fixing conditions
library_construction_protocol: The cells were harvested at 3,000 × g for 2 min at room temperature, and washed by ice-cold phosphate solution (0.1 g K2HPO4 and 0.4 g KH2PO4 in 450 mL distilled water) Wild type and ΔAvin_01720 A. vinelandii strains were cultured on MB plates with a nitrogen source for 2 days, inoculated into MB liquid medium with a nitrogen source for 24 h, and in MB liquid medium without a nitrogen source for 3 h. Then, the vials were sealed and the cells were collected after the additional 2 h incubation. Total RNA was extracted from each culture with TRIzol reagent (Invitrogen, NY, USA). RNA integrity was evaluated for quality control with an Agilent Bioanalyzer 2,100 using Agilent RNA 6,000 Nano Kit (Agilent technologies, CA, USA). After the quality evaluation, rRNAs were removed with RiboMinus Transcriptome Isolation Kit, bacteria (Thermo Fisher Scientific, MA, USA). cDNA library preparation was performed using KAPA RNA HyperPrep Kit Illumina Platforms (Kapa Biosystems, MA, USA). Prepared cDNA libraries were validated with an Agilent Bioanalyzer 2,100 using an Agilent High Sensitivity DNA Kit (Agilent Technologies).