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|Public on Mar 22, 2021
|tissue: peripheral blood
cell type: MAIT
|Isolated cells were fixed with 1% formaldehyde in PBS for 10 minutes, then quenched with glycine. After washing, DNA was extracted using the Illumina SimpleChIP Enzymatic ChIP kit (#9003S). Briefly, nuclei were isolated by incubating in 1ml buffer A for 10 min on Ice, followed by centrifugation and resuspension in 1 ml Buffer B. Chromatin was sheared to roughly 200-1000bp. Samples were centrifuged to clear lysates, and the supernatant containing the DNA was transferred to a fresh tube for overnight antibody complexing. Next day, protein G dynabeads were added to each tube, incubated, washed, and isolated with magnet. DNA complexes were then released from the beads, decrosslinked, and phenol:choloroform:isolamyl alcohol extracted for library prep.
DNA libraries were prepared using NEBNext Ultra II DNA Library Prep Kit for Illumina. Briefly, DNA ends were repaired, followed by ligation of adapter sequences. DNA was cleaned with AMPure XP beads, and then DNA was PCR amplified with 7-8 cycles. After final cleaup with AMPure XP Beads, libraries were quantified using Agilent Bioanalyzer high sensitivity DNA kit and were subsequently sequenced.
|Illumina HiSeq 3000
|CD3+ CD161+ TCR Vα7.2+
|Fastq files prepared using Illumina FASTQ Generation application
Fastq files were trimmed using Trimmomatic
Reads were aligned to the hg19 genome using Bowtie2
Duplicate reads were removed using PicardTools
Peaks were called using Macs2
Supplementary_files_format_and_content: Bedgraph files were generated from Macs2 output
|Feb 23, 2020
|Last update date
|Mar 22, 2021
|Moffitt Cancer Center
|Department of Immunology
|12902 Magnolia Drive, MRC4
|Mapping Bcl11b binding in human MAIT cells
|Bcl11b binding and transcriptome analysis of Bcl11b-deficient MAIT cells at steady state and infection
|SRA Run Selector