|
| Status |
Public on Jan 01, 2020 |
| Title |
taur 2 |
| Sample type |
SRA |
| |
|
| Source name |
ATCC 2511 broth_taurine_DMSO
|
| Organism |
Escherichia coli |
| Characteristics |
strain: K12 MG1655
media: ATCC 2511 modified to have taurine as the sole sulfur source
treatment: 1% DMSO
|
| Treatment protocol |
50 μL of either DMSO or 50 μL of 1.56 mg/mL 2-((1H-1,2,3-triazol-4-yl)thio)-N-(2-fluorophenyl)acetamide was added to a 50 mL conical tube. When E. coli cultures reached mid-log, 5 mL of culture was added to each tube and allowed to incubate with shaking at 37°C and 225 RPM.
|
| Growth protocol |
For each of three bioreplicates, a single colony of Escherichia coli K12 MG1655 was used to inoculate an overnight culture in either ATCC 2511 broth (M9) or the ATCC 2511 media with a molar equivalent of magnesium chloride used to replace magnesium sulfate and taurine used to as the sole sulfur source (M9T). The next day, overnight cultures were diluted 1:100 into fresh M9 or M9T. The cultures were allowed to reach a mid-log OD600 and then treated.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
10 mL RNAprotect Bacteria Reagent was immediately added and mixed with the drugged cultures and pelleted at 5,000XG for 10 minutes. The supernatant was discarded and the pellets were flash frozen. For RNA extraction, pellets of E. coli K12 were re-suspended in 100 µL of 1 mg/mL lysozyme and incubated at 37°C for 10 minutes with 5 seconds vortex every 2 minutes. Subsequent steps were carried out using the RNeasy extraction kit (Qiagen) according to protocol. RNA was eluted in 40-50 µL of supplied elution buffer. Residual genomic DNA contamination was depleted using RNA Clean and Concentrator kit (Zymo Research) following the manufacture’s protocol. Samples were eluted in 50 µL of DNase/Rnase-Free water. Quality of RNA samples were measured using an Agilent 2100 Bioanalyzer and RIN values were all > 7.0.
ScriptSeq™ v2 RNA-Seq Library Preparation Kit, standard protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
1544126_LEE_008_S99_L004
|
| Data processing |
Ribosomal RNA sequences filtered out using BBDuk {https://jgi.doe.gov/data-and-tools/bbtools/}
Reads aligned to the reference Escherichia coli str. K-12 substr. MG1655 genome (NCBI) using Rockhopper {23716638} to generate the gene counts
DESeq2 analysis package {25516281} in the R statistical computing environment used for differential gene expression analysis
Genome_build: GenBank: U00096.3
Supplementary_files_format_and_content: CSV format containing locus tag and read count
|
| |
|
| Submission date |
Dec 17, 2019 |
| Last update date |
Jan 02, 2020 |
| Contact name |
Richard E Lee |
| Organization name |
St. Jude Children's Research Hospital
|
| Department |
Chemical Biology and Therapeutics
|
| Lab |
Richard Lee
|
| Street address |
262 Danny Thomas Place
|
| City |
Memphis |
| State/province |
Tennessee |
| ZIP/Postal code |
38105 |
| Country |
USA |
| |
|
| Platform ID |
GPL25368 |
| Series (1) |
| GSE142164 |
Discovery and characterization of the antimetabolite action of thioacetamide-linked 1,2,3-triazoles as disruptors of cysteine biosynthesis in Gram-negative bacteria |
|
| Relations |
| BioSample |
SAMN13614678 |
| SRA |
SRX7396602 |
