yeaG_Np_2
Escherichia coli K-12
Protocols: Wild-type and ΔyeaG bacteria were grown in Gutnick minimal medium according to (Figueira et al., Sci Rep, 2015), where the sole source of nitrogen is NH4Cl; overnight cultures were grown in medium containing 10 mM NH4Cl and N-limiting growth curves were carried out in medium containing 3 mM NH4Cl. Cultures were grown as in (Figueira et al., Sci Rep, 2015) and sampled at the following time points: N+, when the OD600nm = 0.3; N-, 20 minutes following N runout; and N-24, where bacteria were subjected to 24 hours of N starvation. Two biological replicates were taken for N+ and N-, and three for N-24. Cultures were mixed with a 1:19 phenol:ethanol solution at a ratio of 9:1 culture:solution and then pelleted. Cell pellets were sent to Vertis Biotechnologie AG for downstream processing exactly as previously described by (Sarkar et al., Nucleic Acids Res, 2017). Total RNA was isolated from the cell pellets using a bead mill and mirVana™ RNA isolation kit (Ambion) according to the manufacturer's instructions. Ribosomal RNA (rRNA) from the total RNA sample was depleted using Ribo-Zero rRNA removal kit for bacteria (Epicentre). The rRNA-depleted RNA samples were fragmented using RNase III. Samples were then poly (A)-tailed followed by treatment with Tobacco Acid Pyrophosphatase (TAP, Epicentre). First strand cDNA synthesis was performed using oligo (dT)-adapter primer and M-MLV reverse transcriptase. Resulting cDNAs were PCR amplified. The primers used for PCR amplification were designed for TruSeq sequencing according to the manufacturer's guidelines (Illumina).
ENA first public
2018-06-04
ENA last update
2018-02-09
External Id
SAMEA104584900
INSDC center alias
ICL-CFB
INSDC center name
Imperial College London
INSDC first public
2018-06-04T17:02:50Z
INSDC last update
2018-02-09T14:26:24Z
INSDC status
public
Submitter Id
E-MTAB-6481:yeaG_Np_2
broker name
ArrayExpress
genotype
yeaG
growth condition
Gutnick minimal medium with 3 millimolar NH4Cl
sample name
E-MTAB-6481:yeaG_Np_2
strain
BW25113