|
| Status |
Public on May 06, 2015 |
| Title |
B_S4 |
| Sample type |
SRA |
| |
|
| Source name |
Bacterial cells
|
| Organism |
Bacillus atrophaeus |
| Characteristics |
strain: UCMB-5137
condition: experiment
media: 1C liquid medium with maize root exudate
|
| Treatment protocol |
Bacterial cells were collected for total RNA extraction by mixing with killing buffer that has been previously frozen. Killing buffer was kept on ice during mixing with bacteria culture, 15 ml of the bacteria culture was mixed with 7.5 ml killing buffer. Composition of killing buffer (20 mM Tris-HCl [BDH Laboratory supplies], 5 mM MgCl2[MERCK], 20 mM NaN3[SIGMA], pH 7.5). The mixture was centrifuged at 5,000 rpm for 3 minutes at room temperature. The pellet was washed with 1 ml killing buffer and immediately frozen at -80°C until RNA extraction.
|
| Growth protocol |
Bacterial cultures were inoculated from a frozen stock culture and incubated at 37°C overnight on solid LB medium. A colony from overnight culture was inoculated in 1C medium (with 0.1% glucose) at 24°C for 14 hours with shaking at 180 rpm. An aliquot from the overnight culture was added to a new 1C medium (with 0.1% glucose), in control and supplemented with 0.25 mg/ml maize root exudate in experiment. Composition of 1 C medium (0.7%w/v pancreatic digest of casein [MERCK], 0.3%w/v papain digest of soya flour[MERCK], 0.5%w/v NaCl [MERCK],) 0.1% glucose [MERCK]. Conical flasks with cultures in control and experiment were grown at 24°C for 14.30 h to achieve a transition to stationary phase characterized by culture medium density (or opacity) of OD600= 1.0 units.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated using ZR Fungal/ Bacteria RNA Mini Prep kit from Zymo research Corp. according to manufacturer’s instruction. The concentration and quality of RNA was checked by NanoDrop then Ribolock Ribonuclease inhibitor [Thermo Scientific] was added to prevent RNA degradation.
cDNA was extracted from total RNA and sequenced using MiSeq 500 Illumina platform in Inqaba (Pretoria, South Africa). Library was prepared by MiSeq Library Prep protocol. rRNA reduction per library was performed with Ribo-Zero.
IB MiSeq 500 cycles, approx. 6 GB data
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina MiSeq |
| |
|
| Data processing |
Basecalls performed using IB MiSeq
Using CLC Genomics Workbench 7.0.3 software, MiSeq 500 reads were aligned against DNA sequences of all predicted CDS in the complete genome sequence of Bacillus atrophaues UCMB-5137 (APIW01000000)
Differential gene expression statistics was calculated for experiment-versus-control samples using EDGE, Baggerley's and t-test algorithms implemented in CLC Genomics Workbench 7.0.3
Supplementary_files_format_and_content: Tab separated text file (locus_tag read_count)
|
| |
|
| Submission date |
May 05, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Oleg Reva |
| E-mail(s) |
oleg.reva@up.ac.za
|
| Phone |
+27124205810
|
| Organization name |
University of Pretoria
|
| Department |
Biochemistry
|
| Lab |
Bioinformatics and Computational Biology Unit
|
| Street address |
Lynnwood Rd., Hillcrest
|
| City |
Pretoria |
| ZIP/Postal code |
0002 |
| Country |
South Africa |
| |
|
| Platform ID |
GPL20146 |
| Series (1) |
| GSE68543 |
RNA-Seq gene expression profiling in the plant growth promoting Bacillus atrophaeus UCMB-5137 revealed an alternative strategy of plant colonization compared to Bacillus amyloliquefaciens endophytes |
|
| Relations |
| BioSample |
SAMN03603538 |
| SRA |
SRX1017458 |
