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| Status |
Public on Aug 13, 2019 |
| Title |
ΔnarK ΔNarS + pPL-NarS |
| Sample type |
SRA |
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| Source name |
Bacterial cells
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| Organism |
Salmonella enterica subsp. enterica serovar Typhimurium str. SL1344 |
| Characteristics |
genotype/variation: SL1344 strain with NarS-narK deletion
plasmid: NarS-overexpressed plasmid pPL-NarS
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| Treatment protocol |
Cells were aerobically cultured to an OD 600 of 0.3, filled in 50 ml closed Falcon tubes, and incubated without agitation at 37°C for the indicated times.
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| Growth protocol |
Overnight cultures were grown from a single colony, diluted 1:100 in fresh LB medium, and grown to the OD600 ~ 0.3 at 37°C with shaking at 220 rpm.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Bacterial cultures were mixed with 0.2 (v/v) of stop solution (95% ethanol and 5% phenol) and immediately frozen within liquid nitrogen. Total RNA was isolated using the hot phenol method. Briefly, 2.0 OD600 cell pellet was resuspended with 300 μL lysozyme (#L3790, Sigma-Aldrich) at 0.5 mg/mL, 30 μL 10% SDS and 33 μL 3M sodium acetate (pH5.2). Cleared lysate was mixed with 375 μL saturated phenol (pH4.5) and incubated at 64°C for 6 min with shaking. The aqueous phase was collected after centrifuge and mixed with 375 μL chloroform in Phase Lock Gel tube (#WM5-2302820, TIANGEN Biotech). RNA was precipitated at -80 °C overnight by mixing the aqueous phase (~350 μL) with 700 μL 30:1 ethanol: sodium acetate (pH 6.5) mix. RNA pellets were washed with 80% ethanol and dissolved in ultra-pure water (#10977, Thermo Fisher Scientific), and was quantified using NanoDrop 2000.
Ribosomal RNAs was removed by Biotin-labeled rRNA probe. Purified and fragmentated RNA was then treated by Illumina TruSeq Standard Kit: Fragmented RNA was reverse transcribed to first chain cDNA random oligo and reverse transcriptase. Second chain cDNA was generated with DNA polymerase I and RNase H. dTTP was replaced by dUTP when Second chain cDNA was generated to increase the chain-specificity. Double chain cDNA was then poly A tailed and ligated with adaptor. After PCR application, samples were sequenced in Illumina Hi-seq platform.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
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| Data processing |
SOAP v2.21 (-m 0 -x 1000 -s 28 -l 32 -v 5 -r 1 -p 3) to filter the reads.
HISAT v2.0.1 (beta (-p 8 --phred64 --sensitive --no-discordant --no-mixed -I 1 -X 1000 --rna-strandness RF) for reads mapping.
Bowtie2 v2.2.5 (q --phred64 --sensitive --dpad 0 --gbar 99999999 --mp 1,1 --np 1 --score-min L,0,-0.1 -I 1 -X 1000 --：) for reads mapping.
RSEM：v1.2.12 forward-prob=0.0) to calculate the gene expression levels.
possionDis：(abs log2(FoldChange) >= 1 && FDR <= 0.005) to call for genes with significant difference expression among two samples.
Genome_build: Salmonella enterica subsp. enterica serovar Typhimurium SL1344 (NC_016810.1)
Supplementary_files_format_and_content: *_FPKM.txt: Tab-delimited text files include FPKM values for each Sample.
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| Submission date |
Aug 12, 2019 |
| Last update date |
Aug 14, 2019 |
| Contact name |
Chuan Wang |
| E-mail(s) |
chuanwang@fudan.edu.cn
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| Organization name |
Fudan University
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| Street address |
131 # Dong An Road
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| City |
Shanghai |
| ZIP/Postal code |
200033 |
| Country |
China |
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| Platform ID |
GPL27045 |
| Series (1) |
| GSE135757 |
The conserved 3’ UTR-derived small RNA NarS counteracts the risk of toxic nitric oxide accumulation during nitrate respiration |
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| Relations |
| BioSample |
SAMN12563078 |
| SRA |
SRX6704832 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4027588_pPL-NarS_FPKM.txt.gz |
47.0 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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