GSM2520772: RD42-NI; Deinococcus deserti; RNA-Seq - SRA

SRX2611094: GSM2520772: RD42-NI; Deinococcus deserti; RNA-Seq
1 ILLUMINA (Illumina HiSeq 2000) run: 44M spots, 4.4G bases, 2.8Gb downloads

Submitted by: NCBI (GEO)

Study: Conservation and diversity of the IrrE/DdrO-controlled radiation response in radiation-resistant Deinococcus bacteria

PRJNA377853 • SRP101333 • All experiments • All runs

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The extreme radiation resistance of Deinococcus bacteria requires the radiation-stimulated cleavage of protein DdrO by a specific metalloprotease called IrrE. DdrO is the repressor of a predicted radiation/desiccation response (RDR) regulon, composed of radiation-induced genes having a conserved DNA motif (RDRM) in their promoter regions. Here, we showed that addition of zinc ions to purified apo-IrrE, and short exposure of Deinococcus cells to zinc ions, resulted in cleavage of DdrO in vitro and in vivo, respectively. Binding of IrrE to RDRM-containing DNA or interaction of IrrE with DNA-bound DdrO was not observed. The data are in line with IrrE being a zinc peptidase, and indicate that increased zinc availability, caused by oxidative stress, triggers the in vivo cleavage of DdrO unbound to DNA. Transcriptomics and proteomics of Deinococcus deserti confirmed the IrrE-dependent regulation of predicted RDR regulon genes and also revealed additional members of this regulon. Comparative analysis showed that the RDR regulon is largely well conserved in Deinococcus species, but also showed diversity in the regulon composition. Notably, several RDR genes with an important role in radiation resistance in Deinococcus radiodurans, for example pprA, are not conserved in some other radiation-resistant Deinococcus species. Overall design: Using Illumina HiSeq 2000, RNA-Seq was performed to explore the transcriptome in non-irradiated and irradiated Deinococcus deserti strains lacking gene irrE or ddrOc.

Sample: RD42-NI

SAMN06473341 • SRS2019747 • All experiments • All runs

Organism: Deinococcus deserti

Library:

Instrument: Illumina HiSeq 2000

Strategy: RNA-Seq

Source: TRANSCRIPTOMIC

Selection: cDNA

Layout: SINGLE

Construction protocol: RNAprotect-treated cells were centrifuged and cell pellets stored at -80°C. Cell pellets were pretreated with lysozyme and proteinase K. Total RNAs were isolated using the mirVana RNA isolation kit (Ambion) including DNase treatment. From each RNA preparation (i.e. non-irradiated and irradiated cells), one cDNA synthesis was carried out. RNA was directly poly(A)-tailed using poly(A) polymerase, followed by TAP treatment. An RNA adapter was then ligated to the 5'P of the total RNA samples. First-strand cDNA synthesis was performed using an oligo(dT)-adapter primer and M-MLV H- reverse transcriptase. The resulting cDNAs were PCR-amplified to about 60-90 ng/μl using a high fidelity DNA polymerase. The cDNAs were purified using the Agencourt AMPure XP kit (Beckman Coulter Genomics) and analyzed by capillary electrophoresis. The primers used for PCR amplification were designed for TruSeq sequencing according to the instructions of Illumina, with the 3’-sequencing adapter containing a barcode specific for each library. For Illumina sequencing, the samples were pooled at approximately equimolar amounts and size fractionated in the range between 250 – 500 bp on Agarose Gel. The pools were sequenced on a Illumina HiSeq 2000 machine (read length: 100 bp).

Experiment attributes:

GEO Accession: GSM2520772

Links:

NCBI link: NCBI Entrez (gds)

Runs: 1 run, 44M spots, 4.4G bases, 2.8Gb

Run	# of Spots	# of Bases	Size	Published
SRR5311138	43,980,748	4.4G	2.8Gb	2017-04-03

ID:: 3773991

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