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| Status |
Public on Apr 01, 2021 |
| Title |
Sa_J1074/R2_DbkdR_Val_late_A |
| Sample type |
SRA |
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| Source name |
cell culture grown in CDM with added valine (3mM)
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| Organism |
Streptomyces albidoflavus |
| Characteristics |
strain: J1074/R2
genotype: Delta-bkdR
treatment: 3mM L-valine
time point: production phase (30h after inoculation)
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| Treatment protocol |
The control culture was grown in pure CDM medium. As treatment, 3mM L-valine was added to the sterile medium prior to inoculation. Samples were taken during growth phase (5h after inoculation) and main production phase (20 to 30 hours after inoculation).
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| Growth protocol |
Liquid cultures were incubated in baffled shake flasks (500 mL, 10% filling volume) on an orbital shaker (Multitron, Infors AG, Bottmingen, Switzerland, 5 cm shaking diameter, 230 rpm, 75 % relative humidity), and at 28°C. S. albus was incubated on MS agar at 28 °C for three days until sporulation occurred. Spores of a single colony were collected to inoculate the pre-culture, which was incubated overnight in LB medium. Afterwards, cells were collected (5,000 xg, 25 °C, 6 min), resuspended in main culture medium (per liter: 10g mannitol, 200 mM potassium phosphate buffer (pH 7.8), 15 g (NH4)2SO4, 1 g NaCl, 550 mg MgCl2*7H2O, 200 mg CaCl2, 30 mg 3,4-dihydroxybenzoic acid, 20 mg FeSO4, 2 mg FeCl3*6H2O, 2 mg MnSO4*H2O, 0.5 mg ZnSO4*H2O, 0.2 mg CuCl2*2H2O, 0.2 mg Na2B4O7*10H2O, 0.1 mg (NH4)6Mo7O24*4H2O, 1 mg nicotinamide, 1 mg riboflavin, 0.5 mg thiamine hydrochloride, 0.5 mg pyridoxine hydrochloride, 0.2 mg biotin, and 0.1 mg p-aminobenzoic acid, in addition to 30 g L-1 glass beads (soda-lime glass, 5 mm, Sigma-Aldrich) to avoid cell agglomeration), and used to inoculate the main culture.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using the Quick-RNA Miniprep Plus kit (Zymo Research, Freiburg, Germany). After DNase treatment, the obtained RNA was purified (RNA Clean & Concentrator-5 kit, Zymo Research) and quantified (DropSense 16, Trinean NV, Gent, Belgium).
To construct whole transcriptome cDNA libraries, 2.5 μg of total RNA each was depleted of rRNA (Ribo-Zero rRNA Removal Kit (Bacteria), Illumina, San Diego, CA, USA). Successful rRNA removal was validated (Agilent RNA Pico 6000 kit, Agilent 2100 Bioanalyzer, Agilent Technologies). The obtained mRNA was converted into a cDNA library according to the TruSeq Stranded mRNA Sample Preparation guide (Illumina). Appropriate cDNA quality and quantity was validated (Agilent High Sensitivity DNA kit, Agilent 2100 Bioanalyzer, Agilent Technologies). Sequencing was performed on an Illumina NextSeq 500 instrument using 75 bases read length (Illumina).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
DbkdR_Val_late_A
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| Data processing |
Basecalls were performed using RTA v 1.18.66.3.
Sequenced reads were mapped to the CP071123.1 whole genome using bowtie2 v2.3.2 with parameters -X 600.
Raw read counts per gene were calculated using featureCounts v2.0.0 with parameters -O -M -t gene -g locus_tag -s 1.
Read count normalization and data analysis were performed using DESeq2.
Genome_build: CP059254.1
Supplementary_files_format_and_content: *.txt: Tab-delimited text files contain normalized read counts derived via DESeq2 for each gene.
Supplementary_files_format_and_content: Raw_Read_Count_Matrix.txt: Tab-delimited matrix table with raw read counts for every gene and every sample.
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| Submission date |
Mar 09, 2021 |
| Last update date |
Apr 02, 2021 |
| Contact name |
Christian Ruckert-Reed |
| E-mail(s) |
[email protected]
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| Phone |
+49 521 106 86308
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| Organization name |
Bielefeld University
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| Department |
Medical School OWL
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| Lab |
CF Omics NGS
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| Street address |
Sequenz 1
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| City |
Bielefeld |
| ZIP/Postal code |
33615 |
| Country |
Germany |
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| Platform ID |
GPL29830 |
| Series (1) |
| GSE168592 |
Superior production of heavy pamamycin derivatives using a bkdR null mutant of Streptomyces albus J1074/R2 |
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| Relations |
| BioSample |
SAMN18234750 |
| SRA |
SRX10298368 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5150287_DbkdR_Val_late_A.txt.gz |
34.2 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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