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| Status |
Public on Aug 11, 2022 |
| Title |
Term-Seq dAR-2 |
| Sample type |
SRA |
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| Source name |
bacterial cell culture
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| Organism |
Bacillus subtilis |
| Characteristics |
genotype: rpoC-10His amyE::Physpank-nusA lacIq -delta-nusA rho::tet Cmr Spr Emr Tetr
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| Treatment protocol |
None
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| Growth protocol |
cells were plated on LB plates at 37°C w/ +2 mM IPTG when required and +20 µg/ml chloramphenicol -> single colonies isolated and grown O/N at 37°C standing culture in +0.4 mM IPTG when required and +20 µg/ml chloramphenicol -> next day, cells were grown at in LB at 37°C +20 µg/ml chloramphenicol
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from collected cell paste via RNeasy mini-kit (Qiagen, cat # 74104)
total RNA was dephosphorylated using CIP (NEB, discontinued), rRNA was depleted via ribo-zero Gram-positive rRNA depletion kits (Illumina, discontinued) or MICROBExpress (Thermo Fisher), remaining RNA was ligated to custom unique 2', 3’ dideoxy RNA oligonucleotide (IDT), Illumina sequencing libraries were generated from ligation products using TruSeq Stranded mRNA kit (Illumina, cat # 20020594) or NEBNext Ultra Directional (NEB, cat # E7760L)
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
dAR-2
NusA depletion rho deletion - Termseq
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| Data processing |
Library strategy: Term-seq
base calling and quality scoring conducted via BaseSpace
adaptor trimming and low quality read removal via Trimmomatic v. 0.38 for RNA-seq .fastq dataset
all reads containing custom oligo were extracted via Cutadapt v. 1.16 for Term-seq .fastq dataset
both RNA-seq and Term-seq .fastq files were mapped to B. subtilis genome using bwa-mem v.0.7.12-r1034
sam -> bam, and bam files split by strand via samtools v. 0.1.19-44428cd
coverage files (.cov) for 3' end mapping and .bedgraph files to illustrate coverage were generated using bedtools v. 2.26.0
2 custom python (python v. 3.8.3) scripts were used to identify 3' ends from Term-seq .fastq derived .cov files (located at: https://github.com/zfmandell/Term-seq)
RNA-seq coverage files generated using bedtools v. 2.26.0
Genome_build: Refseq : NC_000964.3
Supplementary_files_format_and_content: bedgraph files, containing a list of called 3' ends with associated height values
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| Submission date |
Nov 07, 2021 |
| Last update date |
Aug 12, 2022 |
| Contact name |
Paul Babitzke |
| E-mail(s) |
pxb28@psu.edu
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| Organization name |
Pennsylvania State University
|
| Street address |
203 Althouse Laboratory
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| City |
State College |
| State/province |
PA |
| ZIP/Postal code |
16802 |
| Country |
USA |
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| Platform ID |
GPL30886 |
| Series (1) |
| GSE188366 |
Rho stimulates both Rho-dependent and intrinsic termination in Bacillus subtilis |
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| Relations |
| BioSample |
SAMN22964129 |
| SRA |
SRX13037143 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5678301_AR_2_Term-ends.bedgraph.gz |
56.5 Kb |
(ftp)(http) |
BEDGRAPH |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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