|
| Status |
Public on Sep 01, 2018 |
| Title |
SL476 control [SHC331_C] |
| Sample type |
SRA |
| |
|
| Source name |
SL476 control
|
| Organism |
Salmonella enterica subsp. enterica serovar Heidelberg |
| Characteristics |
isolate: SL476
antimicrobial resistance: Gen Amp Kan Str Tet Aug Cep Smx
pfge pattern: Unknown
source type: Food isolate; reference strain
culture condition: Stationary-phase culture grown at 37C
|
| Biomaterial provider |
Salmonella Genetic Stock Center, Calgary
|
| Treatment protocol |
2mL stationary-phase samples were heat shocked at 56C for 7.5 minutes, with a time-to-temperature of 2.43 minutes. A 10% ice-cold phenol:ethanol stop-solution was immediately added to each solution upon removal from heat and samples were placed on ice
|
| Growth protocol |
Overnight aerobic cultures in tryptic soy broth (TSB,BD, Franklin Lake, NJ) started from a single colony on tryptic soy agar (TSA; BD, Franklin Lake, NJ) were diluted to 103 CFU/mL into TSB and grown for 12 hours at 37C, 200 rpm to stationary phase (10 log10 CFU/mL) prior to RNA extraction. Growth phase had been previously confirmed by dilution plating onto TSA.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted using Trizol (Ambion, Foster City, CA) with an additional phenol:chloroform extraction in triplicate for each isolate and condition. RiboZero (Illumina, San Diego, CA) was used to remove rRNA; TURBO DNAse (Thermo Fisher, Waltham, MA) was used to remove DNA. DNA removal was assessed using qRT-PCR for rpoD and gyrB (Table 3.2) (25, 34, 61, 80). Primers and probes were purchased from Integrated DNA Technologies (IDT, Coralville, IA); qRT-PCR was performed using IDT’s 2x master mix. If negative control samples (RNA samples with no reverse transcriptase added) did not amplify across the CT threshold prior to cycle 36, the sample was deemed free of DNA contamination.
Illumina's ScriptSeq v. 2 bacterial kit with Ribozero gold; constructed with low-concentration protocol
Enzymatic fragmentation, random-hexamer priming for amplification
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
File names with C indicate control samples, samples with H indicate heat shock samples
022013_SHC-3-31_ATCACG
|
| Data processing |
samples quality trimmed by sequencing core to Q20 using Trimmomatics; adaptors and barcodes also removed in this step
SL476 index constructed from NCBI reference sequence (NC_011083.1) using Bowtie 2 (v. 2.1.0)
Trimmed reads were aligned with the SL476 index using Tophat (v.2.1.1)
Tophat Bam files were sorted using sam tools to obtain name-sorted .sam files necessary for HTSeq proper functioning
Read counts were obtained using HTSeq (v.0.6.1)
Genome_build: SL476 refseq genome NC_011083.1
Supplementary_files_format_and_content: Name-sorted read count files (Tophat 2 output). In tab-delimited text file format which can be opened using Excel or a text editor. Each file contains a column with the gene name and the number of reads associated with that gene
|
| |
|
| Submission date |
Sep 01, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Andrea J Ray |
| E-mail(s) |
[email protected]
|
| Organization name |
Purdue University
|
| Department |
Food Science
|
| Lab |
Oliver Lab
|
| Street address |
745 Agriculture Mall Drive
|
| City |
West Lafayette |
| State/province |
IN |
| ZIP/Postal code |
47907 |
| Country |
USA |
| |
|
| Platform ID |
GPL23979 |
| Series (1) |
| GSE103418 |
Comparison of gene expression profiles for heat-shocked and non-heat shocked stationary phase samples from heat tolerant isolates R1-0006 and R1-0007 and reference strain SL476 |
|
| Relations |
| BioSample |
SAMN07599834 |
| SRA |
SRX3156449 |
