|
| Status |
Public on Aug 26, 2021 |
| Title |
C1B |
| Sample type |
SRA |
| |
|
| Source name |
Mtb phosphate limited (RegX3-IP) rep2
|
| Organism |
Mycobacterium tuberculosis |
| Characteristics |
media: phosphate limited
chip antibody: anti-RegX3 (custom-made antibody of RegX3 was raised by Thermo Fisher Scientific, USA)
molecule subtype: IP DNA
|
| Treatment protocol |
Log phase cells were harvested and resuspended in phosphate limited medium and grown for 72 hours under shaking at 37◦C
|
| Growth protocol |
Mtb was grown to log phase in Middle Brook 7H9 medium containing 10% ADC
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Mtb grown in phosphate depleted medium was crosslinked using 1% formaldehyde followed by quenching with 250mM glycine. The cells were lysed and the DNA was sheared on Bioruptor plus (Diagenode) with 25 cycles of 30 sec/ 90 sec on/off, respectively, so as to generate fragment sizes of ~250 bp. Immune complexes were precipitated with RegX3 antibody or control IgG, and washed. DNA from total lysates (input) and immunoprecipitates (IP) were purified after decrosslinking with Proteinase K and samples were used immediately for next-generation sequencing (NGS).
ChIP-Seq libraries for sequencing were constructed at Genotypic Technology’s Genomics facility according to the NEXTflex ChIP-Seq library protocol outlined in the NEXTflex ChIP-Seq kit (cat nos102–1001)
The libraries were enriched using PCR followed by purification using Agencourt AMPURE XP beads (Beckman Coulter #A63881) and quality was checked on a High Sensitivity Bioanalyzer Chip (Agilent).Sequencing was performed at Genotypic Technology’s Genomics facility following certified protocols from Illumina on an Illumina Hi-seq platform using single-end 75 bp chemistry.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
C1B
|
| Data processing |
The quality check of raw reads was performed using the Genotypic FastQC tool.
The reads were aligned to the reference M. tuberculosis H37Rv genome using the Bowtie alignment tool
Peak detection was carried out using the Hypergeometric Optimization of Motif EnRichment (Homer) tool.
Genome_build: H37Rv
|
| |
|
| Submission date |
Aug 24, 2021 |
| Last update date |
Aug 27, 2021 |
| Contact name |
Manikuntala Kundu |
| E-mail(s) |
manikuntala.kundu@gmail.com
|
| Organization name |
Bose Institute
|
| Street address |
93/1, Acharya Prafulla Chandra Road
|
| City |
Kolkata |
| State/province |
West Bengal |
| ZIP/Postal code |
700009 |
| Country |
India |
| |
|
| Platform ID |
GPL22688 |
| Series (1) |
| GSE182669 |
Genome-wide association study of RegX3 in M. tuberculosis (Mtb) grown under phosphate limited conditions |
|
| Relations |
| BioSample |
SAMN20959931 |
| SRA |
SRX11895551 |
