|
| Status |
Public on Dec 16, 2021 |
| Title |
CompD1 |
| Sample type |
SRA |
| |
|
| Source name |
Mycobacterium tuberculosis bacteria
|
| Organism |
Mycobacterium tuberculosis CDC1551 |
| Characteristics |
genotype: Rv1625c Complement
compound treatment: DMSO (Vehicle)
|
| Growth protocol |
Bacterial strains were pre-grown in 7H9OADC before inoculation into fresh media containing a cAMP-inducing compound (10 µM V-59 or 500 ng/mL Atc) or vehicle control (DMSO or EtOH) at an OD600 of 0.1. The following day, bacteria were inoculated into 7H12+cholesterol media containing fresh compound or vehicle control. After four hours, cultures were pelleted by centrifugation, washed with guanidine thiocyanate-based buffer and stored at -80°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol LS (Thermo Fisher). Pellets were washed and suspended in Trizol LS (Ambion), 0.1 mmol silicon beads were added, and cells were lysed using a BeadBeater. Total RNA was isolated by chloroform extraction and precipitated in isopropanol with GlycoBlue reagent (Thermo Fisher). RNA was resuspended in nuclease-free water, genomic DNA contamination was removed using the Turbo-DNA free kit (Invitrogen), and rRNA was depleted using the Ribo-Zero Gold rRNA removal kit (Illumina).
Sample quality was determined via Fragment Analyzer (Advanced Analytical) and TruSeq-barcoded RNAseq libraries were generated with the NEBNext Ultra II Directional RNA Library Prep Kit (New England Biolabs). Sequencing was performed at the Cornell University Transcriptional Regulation and Expression Facility on a NextSeq500 instrument (Illumina) at a depth of 15 M single-end 75 bp reads.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
11236_10135_108921_H7Y3TBGXC_1092-3319-KW13-CompD_1_CATTTT_R1.fastq.gz
|
| Data processing |
Basecalling was perfomed on the Illumina NextSeq 500 using RTA 2.0.
Reads were trimmed for low quality and adaptor sequences with TrimGalore v0.6.0, a wrapper for cutadapt and fastQC. Parameters: -j 1 -e 0.1 --nextseq-trim=20 -O 1 -a AGATCGGAAGAGC --length 50 --fastqc
Reads were mapped to the Mycobacterium tuberculosis CDC1551 reference genome using STAR v2.7.0e. Parameters: --outSAMstrandField intronMotif , --outFilterIntronMotifs RemoveNoncanonical ,--outSAMtype BAM SortedByCoordinate, --quantMode GeneCounts
SARTools and DESeq2 v1.26.0 were used to generate normalized counts and statistical analysis of differential gene expression, upon which conclusions were based.
Genome_build: Mycobacterium tuberculosis CDC1551 reference genome (GCA_000008585.1 )
Supplementary_files_format_and_content: tab-delimited text file containing normalized counts for each sample and each gene
|
| |
|
| Submission date |
Dec 14, 2021 |
| Last update date |
Dec 16, 2021 |
| Contact name |
Brian VanderVen |
| E-mail(s) |
bcv8@cornell.edu
|
| Organization name |
Cornell University
|
| Department |
Microbiology and Immunology
|
| Street address |
930 Campus Road
|
| City |
Ithaca |
| State/province |
New York |
| ZIP/Postal code |
14850 |
| Country |
USA |
| |
|
| Platform ID |
GPL31090 |
| Series (1) |
| GSE190875 |
Transcriptional changes associated with chemical induction of adenylyl cyclase activity in Mtb |
|
| Relations |
| BioSample |
SAMN24022225 |
| SRA |
SRX13406868 |
