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| Status |
Public on Aug 07, 2021 |
| Title |
Lbr35-minus-MSG-1-T12 |
| Sample type |
SRA |
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| Source name |
bacterial pellet
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| Organism |
Levilactobacillus brevis |
| Characteristics |
strain: Lbr35
msg_condition: minus
timepoint: 12
replicate: 1
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| Treatment protocol |
MRS media +/- MSG
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| Growth protocol |
Overnight cultures (16-18 h) of L. brevis Lbr-6108, and Lbr-35 were inoculated in 15 mL disposable culture tubes (p/n 14-961-27; Fisher, Hanover Park, IL, USA) containing 3 mL of media to reach an optical density (OD) of approximately 0.100 measured at 600 nm using a spectrophotometer (Genesys 20; Thermo Fisher, Waltham, MA, USA). For each strain, two types of media were used; MRS broth (BD) without any other supplementation (further denoted as MRS) and MRS broth supplemented with 10,000 μg/mL (equivalent to 10 g/L) of L(+)-glutamic acid monosodium salt monohydrate (p/n 119940010; Acros Organics, Morris Plains, NJ, USA) (further denoted as MRS+MSG). The OD measurement for timepoint zero (T0) was measured shortly after inoculation of the overnight culture into disposable culture tubes containing 3 mL media. Cultures for all except ATCC 14869 were harvested at the initial timepoint (T0) and then every 3 h post-inoculation until 24 h, and then only at 48 h post-inoculation and the OD was measured just before harvesting. For ATCC 14869, the cultures were harvested after measuring the OD at 9 h, 24 h and then at 48 h post-inoculation because of its slower growth pattern. For each harvest timepoint separate sets of culture tubes were prepared, so that only the set of tubes for the designated time points were removed from the incubator for harvesting, without disturbing the culture growth. All culture tubes with strains growing in MRS or MRS+MSG were placed in the 37°C ± 1°C incubator under anaerobic conditions using anaerobic gas packsGasPak EZ container systems (p/n 260001, BD). Samples were removed from the incubator at respective timepoints and the ODs were measured at 600 nm.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA from Lbr-6018 and Lbr-35 RNA were extracted from cell pellets in RNA protect at GENEWIZ
Ribosomal RNA depletion was performed using FastSelect bacteria reagents (Qiagen, Hilden, Germany). RNA sequencing library preparation used NEBNext Ultra II RNA Library Preparation Kit for Illumina by following the manufacturer’s recommendations (NEB, Ipswich, MA, USA). Briefly, enriched RNAs are fragmented for 15 minutes at 94 °C. First strand and second strand cDNA are subsequently synthesized. cDNA fragments are end repaired and adenylated at 3’ends, and universal adapters are ligated to cDNA fragments, followed by index addition and library enrichment with limited cycle PCR.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
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| Description |
total RNA extracted from bacterial culture pellet
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| Data processing |
RTA v. 2.7.7 was used for basecalling, and bcl2fastq-v2.20.0 was used for demultiplexing
Reads were trimmed with trimgalore using default settings, and then transcripts were quantified using salmon v 1.3.0 with the respective transcriptome generated from the genome and gff file.
Transcript quantifications were read in to R using tximport v 1.14.2, and normalized with DESeq2 v 1.26.0 note: we didn't end up processing the T48 timepoint samples due to quality issues, but we included them in case others would like to have access to the data.
Genome_build: Lbr-35 = JAHERW000000000
Genome_build: Lbr-6108 = MDUA01000000
Supplementary_files_format_and_content: matrix table of normalized gene abundance for each sample and each gene
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| Submission date |
Aug 05, 2021 |
| Last update date |
Aug 07, 2021 |
| Contact name |
Bryan Zabel |
| E-mail(s) |
bryan.zabel@iff.com
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| Organization name |
International Flavors & Fragrances Inc
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| Department |
GEM
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| Street address |
3329 Agriculture Dr
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| City |
Madison |
| State/province |
wi |
| ZIP/Postal code |
53716 |
| Country |
USA |
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| Platform ID |
GPL30477 |
| Series (1) |
| GSE181504 |
Transcriptomics reveal strain-specific metabolic strategies for acid resistance and gamma-aminobutyric acid (GABA) production in Levilactobacillus brevis. |
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| Relations |
| BioSample |
SAMN20592052 |
| SRA |
SRX11652082 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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