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| Status |
Public on Jun 01, 2020 |
| Title |
AAP5, high carbon, dark, Replicate 2 |
| Sample type |
SRA |
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| Source name |
AAP5 cultivated in liquid organic medium
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| Organism |
Sphingomonas sp. AAP5 |
| Characteristics |
treatment: growth in full medium, harvested in dark
time (day): 2
replicate: 2
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
For isolation and purification the RNeasy-kit (Qiagen) was used according to the manufacturer’s manual. Modifications are listed below. Cells lysis was performed enzymatically using 15 mg/mL lysozyme in Tris-EDTA-buffer (ph 8.0) and mechanically by vortexing for 3 min with acid washed glass beads. A first digestion of genomic DNA was performed on the column, using RNase-free DNase I (Qiagen) according to the manufacturer’s protocol. Total RNA was eluted in 88 µL RNase-free H2O and a second DNase I-digestion of genomic DNA was performed in solution, followed by a second RNeasy purification step. In this second purification an additional washing step with 80 % ethanol was performed prior to elution with 30µl RNase-free water. The concentration of the RNA was quantified using a NanoDrop spectrophotometer (Peqlab, Erlangen, Germany) and the RNA integrity was assessed using Bioanalyzer 2100 (Agilent, Santa Clara, USA). Ribosomal RNA depletion: The Ribo-Zero (gram-negative) kit to remove the prokaryotic rRNA, according to the manufacturer’s instructions.
Single end, strand specific cDNA libraries were prepared using the Scriptseq v2 RNA-Seq Library Preparation Kit (Illumina, San Diego, CA, USA) following the manufacturers protocol. For sequencing equal volumes of libraries (12 pM) were multiplexed on a single lane. Sequencing was done on the HiSeq 2500 (Illumina, San Diego, CA, USA) using TruSeq SBS Kit v3 - HS (Illumina, San Diego, CA, USA) for 50 cycles resulting in 50 bp reads. Image analysis and base calling were performed using the Illumina pipeline v 1.8 (Illumina, San Diego, CA, USA).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
D1-2
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| Data processing |
The demultiplexed raw fastq-files were quality-controlled using the FASTQ-mcf suite (https://github.com/ExpressionAnalysis/ea-utils). Low quality bases (Phred-score < 30) and Illumina adaptors were clipped. Reads were mapped to the reference genome using bowtie2 (Langmead et al., 2012) with default parameters for single end reads. Ambiguously mapping reads were randomly distributed between all regions to which they could be assigned. The resulting sam-files were converted to indexed binary format and pile-up format using samtools (Li et al., 2009). The program featureCounts (Liao et al., 2014) was used to count the reads mapping to genes.
Genome_build: NZ_CP037913.1, NZ_CP037915.1, NZ_CP037914.1, NZ_CP037916.1
Supplementary_files_format_and_content: tab-delimited text file contains reads per gene
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| Submission date |
Mar 16, 2020 |
| Last update date |
Jun 02, 2020 |
| Contact name |
Karel Vaclav Kopejtka |
| E-mail(s) |
kopejk00@alga.cz
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| Phone |
+420777623254
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| Organization name |
Institute of Microbiology Czech Acad. Sci.
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| Department |
Centre Algatech
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| Lab |
Laboratory of Anoxygenic Phototrophs
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| Street address |
Novohradska
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| City |
Trebon |
| ZIP/Postal code |
37981 |
| Country |
Czech Republic |
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| Platform ID |
GPL28277 |
| Series (2) |
| GSE147051 |
Transcriptome response to carbon-limiting conditions. |
| GSE147102 |
Simultaneus presence of bacteriochlorophyll and xanthorhodopsin genes in freshwater bacterium Sphingomonas sp. AAP5 |
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| Relations |
| BioSample |
SAMN14383857 |
| SRA |
SRX7917671 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4413702_D1-2_S32_L004_R1_001_sorted_count.txt.gz |
60.6 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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