|
| Status |
Public on Aug 16, 2018 |
| Title |
4_h_MMC_minusTEX |
| Sample type |
SRA |
| |
|
| Source name |
4_h_MMC_total bacterial RNA
|
| Organism |
Caulobacter vibrioides NA1000 |
| Characteristics |
genotype/variation: wild type
internal stock id: KFS_0006
treated with: 0.1 ug/mL mitomycinC (MMC) for 4hrs
|
| Treatment protocol |
Samples were collected prior to treatment (OD660 of 0.4) and after 4h of treatment with mitomycinC (0.1 ug/mL)
|
| Growth protocol |
Caulobacter crescentus was grown aerobically at 30C in PYE medium
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted with the HotPhenol method and digested with DNaseI
For the depletion of processed transcripts, equal amounts of RNA were incubated with Terminator 5'-phosphate-dependent exonuclease (TEX) (Epicentre #TER51020) as previously described (Sharma et al., 2010). The transcripts were not fragmented in order to get mainly sequencing reads of the 5'-end of the transcripts. Libraries for Illumina sequencing of cDNA were constructed by vertis Biotechnology AG, Germany (http://www.vertis-biotech.com/), as described previously for eukaryotic microRNAs (Berezikov et al., 2006) but omitting the RNA size-fractionation step prior to cDNA synthesis. Equal amounts of RNA samples were poly(A)-tailed using poly(A) polymerase. Then, the 5'-triphosphates were removed by applying tobacco acid pyrophosphatase (TAP) resulting in 5'-monophosphat. Afterwards, a RNA adapter was ligated to the 5'-phosphate of the RNA. First-strand cDNA was synthesized by an oligo(dT)-adapter primer and the M-MLV reverse transcriptase. In a PCR-based amplification step using a high fidelity DNA polymerase the cDNA concentration was increased to 20-30 ng/µl. A library-specific barcode for multiplex sequencing was part of a 3'-sequencing adapter.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Quality trimming with fastq_quality_trimmer (fastx 0.11.2)
Read mapping with READemption (0.4.3) and segemehl (0.2.)
Coverage plote generation with READemtpoin
Genome_build: NC_011916.1
Supplementary_files_format_and_content: wiggle of normalized coverages
|
| |
|
| Submission date |
Sep 25, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Konrad U. Förstner |
| E-mail(s) |
foerstner@zbmed.de
|
| Organization name |
ZB MED - Information Centre for Life Sciences
|
| Department |
Information Services
|
| Lab |
Förstner Lab
|
| Street address |
Gleueler Str. 60
|
| City |
Cologne |
| State/province |
North Rhine-Westphalia |
| ZIP/Postal code |
50931 |
| Country |
Germany |
| |
|
| Platform ID |
GPL21317 |
| Series (1) |
| GSE104186 |
Identification of small RNAs expressed in Caulobacter crescentus in response to DNA damage |
|
| Relations |
| BioSample |
SAMN07692732 |
| SRA |
SRX3210366 |
