|
| Status |
Public on Jan 23, 2019 |
| Title |
0_05-1 |
| Sample type |
SRA |
| |
|
| Source name |
E. coli bacteria + P. putida bacteria + IncPα plasmid
|
| Organisms |
Pseudomonas putida; Escherichia coli |
| Characteristics |
e. coli strain: K-12 LE392
p. putida strain: KT2440
plasmid: IncPalpha RP4 plasmid
protocol: 0.05 mg/L carbamazepine
|
| Treatment protocol |
Mixed wild-type strains E. coli K-12 LE392, P. putida KT2440 and IncPα RP4 plasmid were cultured in triplicate in 10 mL 0, 0.05, 10.0, and 50.0 mg/L carbamazepine, at 25 °C, without shaking
|
| Growth protocol |
liquid LB culture at 30 °C
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Bacterial cells were then harvested by centrifugation at 8000 × g for 10 min. Total RNA was extracted from the mutants using the QIAGEN miRNeasy Mini Kit (QIAGEN, Germany) manufacturer’s protocol with one extra bead-beating step to completely lyse the bacterial cells.
RNA libraries were prepared for sequencing using standard Illumina protocols
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Base calling through an integrated primary analysis software called RTA (Real Time Analysis. v1.18).
The NGS QC toolkit (version 2.3.3) was used to treat the raw sequence reads to trim the 3’-end residual adaptors and primers, and the ambiguous characters in the reads were removed. Then, the sequence reads consisting of at least 85% bases were progressively trimmed at the 3’-ends until a quality value ≥ 20 were kept. Downstream analyses were performed using the generated clean reads of no shorter than 75 bp.
The clean reads of each sample were aligned to the E. coli reference genome (NC_000913), P.putida reference genome (NC_002947), and IncPα plasmid reference genome (NC_00) using SeqAlto (version 0.5). Cufflinks (version 2.2.1) was used to calculate the strand-specific coverage for each gene, and to analyze the differential expression in triplicate bacterial cell cultures. The statistical analyses and visualization were conducted using CummeRbund package in R (http://compbio.mit.edu/cummeRbund/).
Gene expression was calculated as fragments per kilobase of a gene per million mapped reads (FPKM, a normalized value generated from the frequency of detection and the length of a given gene.
Genome_build: NC_000913.3 NC_002947.4 L27758.1
Supplementary_files_format_and_content: Excel files include FPKM values for each sample
|
| |
|
| Submission date |
Mar 20, 2018 |
| Last update date |
Jan 23, 2019 |
| Contact name |
Jianhua Guo |
| E-mail(s) |
j.guo@awmc.uq.edu.au
|
| Organization name |
University of Queensland
|
| Department |
Australian Centre for Water and Environmental Biotechnology
|
| Street address |
Research Road, Level 4 Gehrmann Building
|
| City |
Brisbane |
| ZIP/Postal code |
4072 |
| Country |
Australia |
| |
|
| Platform ID |
GPL24748 |
| Series (1) |
| GSE112064 |
Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and carbamazepine-stressed E. coli K-12 LE392, P. putida KT2440, and RP4 plasmid Transcriptomes |
|
| Relations |
| BioSample |
SAMN08742151 |
| SRA |
SRX3823276 |
