|
| Status |
Public on Jun 07, 2023 |
| Title |
WT PYE + 9.3 mM gln log phase - rep 3 |
| Sample type |
SRA |
| |
|
| Source name |
cultures
|
| Organism |
Caulobacter vibrioides NA1000 |
| Characteristics |
tissue: cultures
treatment: 9.3 mM (final concentration) glutamine
|
| Treatment protocol |
For glutamine treatment conditions, 9.3 mM (final concentration) glutamine was added to growth medium
|
| Growth protocol |
Starter cultures were grown for 18 hours at 30˚C in PYE or PYE plus 9.3mM (final concentration) glutamine. Cultures were then diluted to OD660 0.1 in their respective medium and grown for 2 h to get the cultures in similar (logarithmic) phase of growth. Once again, cultures were diluted to OD660 0.1 in their respective medium and grown another 3.25 h (OD660 < 0.4) to capture mRNA in similar log-phase growth.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
6 ml of each culture were pelleted via centrifugation (1 min at 17,000 x g). Pellets were immediately resuspended in 1ml TRIzol and stored at -80˚C until RNA extraction. To extract RNA, thawed samples were incubated at 65˚C for 10 min. After addition of 200 µl of chloroform, samples were vortexed for 20 s and incubated at RT for 5 min. Phases were separated by centrifugation (10 min at 17,000 x g). The aqueous phase was transferred to a fresh tube and an equal volume of isopropanol was added to precipitate the nucleic acid. Samples were stored at 80˚C (1 h to overnight), then thawed and centrifuged at 17,000 x g for 30 min at 4˚C to pellet the nucleic acid. Pellets were washed with ice-cold 70% ethanol then centrifuged for at 17,000 x g for 5 min at 4˚C. After discarding the supernatant, pellets were air-dried at RT, resuspended in 100 µl RNAse-free water, and incubated at 60˚C for 10 min. Samples were treated with TURBO DNAse (Invitrogen) following manufactures protocol for 30 min at RT and then column purified using RNeasy Mini Kit (Qiagen). RNA samples were sequenced at SeqCenter (Pittsburgh, PA). Briefly, sequencing libraries were prepared using Illumina’s Stranded Total RNA Prep Ligation with Ribo-Zero Plus kit and custom rRNA depletion probes. 50 bp paired end reads were generated using the Illumina NextSeq 2000 platform (Illumina).
Library preparation was performed using Illumina’s Stranded Total RNA Prep Ligation with Ribo-Zero Plus kit and 10bp IDT for Illumina indices
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
NextSeq 2000 |
| |
|
| Data processing |
Reads were mapped to NA1000 Caulobacter crescentus genome and normalized using default settings for RNA-seq workflow in CLC Genomics Workbench v 21
Assembly: NA1000 Caulobacter crescentus
Supplementary files format and content: processed data file contains normalized counts (counts per million (CPM)) for each annotated gene in NA1000 Caulobacter crescentus genome
|
| |
|
| Submission date |
Jun 05, 2023 |
| Last update date |
Jun 07, 2023 |
| Contact name |
Sean Crosson |
| E-mail(s) |
crosson4@msu.edu
|
| Phone |
5178845345
|
| Organization name |
Michigan State University
|
| Department |
Dept. Microbiology and Molecular Genetics
|
| Street address |
567 Wilson Rd
|
| City |
East Lansing |
| State/province |
Michigan |
| ZIP/Postal code |
48824 |
| Country |
USA |
| |
|
| Platform ID |
GPL33463 |
| Series (2) |
| GSE234095 |
The Caulobacter NtrB-NtrC two-component system bridges nitrogen assimilation and cell development [RNA-seq] |
| GSE234097 |
The Caulobacter NtrB-NtrC two-component system bridges nitrogen assimilation and cell development |
|
| Relations |
| BioSample |
SAMN35621719 |
| SRA |
SRX20592344 |
