|
| Status |
Public on Mar 08, 2021 |
| Title |
E14-5 whole lung NKX2-1 ChIP-seq rep 2 |
| Sample type |
SRA |
| |
|
| Source name |
Lung tissue
|
| Organism |
Mus musculus |
| Characteristics |
chip antibody: NKX2-1 (ab133737, Abcam)
age: E14-5
cell type: whole lung
replicate: 2
|
| Treatment protocol |
YT Sox9 mutants were induced at E15.5 with 3mg of Tamoxifen, GFP labeling of AT2 cells was carried out by 250ug of Tamoxifen at P4 for P7 AT2 cells and 3mg of Tamoxifen at 10 wks, 3 days prior harvest the adult.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Whole lung ChIP-seq were performed as described in the associated paper and cell type specific ChIp-seqs were performed on chromatin from sorted nulcei based on a SunGFP reporter induced by recombination undercontrol of cell type specific Cre or CreERs.
ChIP DNA quantity was measured using the Qubit dsDNA HS Assay Kit (Invitrogen, Q32851). 2-5ng of sample DNA was used to generate sequencing libraries using the NEB Next Ultra II DNA Library Prep Kit for Illumina (NEB, E7645). Step 3.1 in the NEB protocol was skipped to improve sample retention per manufacturer’s recommendation. Samples were PCR amplified for 12 cycles and barcoded using indexing primers (New England BioLabs, E7335s or E7500S) followed by a final (.65 x -1 x bead volume) size selection and purification step using a SPRIselect reagent (Beckman Coulter, B23318). Samples were then verified for library size by gel electrophoresis and concentration measured using the Qubit HS dsDNA assay. Samples were then sequenced on the Illumina NextSeq500.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Fastqc was run on the raw Fastq files to determine initial quality of reads.
Trimmomatic was performed to remove barcodes and poor quality bases.
Fastqc was run for quality control.
Bowtie alignment to the mm10 genomic with -m1 -k1 -v1 parameters and filtered for unmapped reads with sam file output
After alignment, sam files were converted to bam files and filtered for unmapped reads, chimeric alignments, low quality alignments, and PCR duplicates using Picard59 MarkDuplicates and samtools60 settings: -b -h -F 4 -F 1024 -F 2048 -q 30.
Peaks were then called using MACS263,64 with the default settings for NKX2-1, H3K4me3, and H3K27ac, and the –broad setting for the broad histone mark H3K4me1.
Genome_build: mm10
Supplementary_files_format_and_content: bedgraph
Supplementary_files_format_and_content: peaks.narrowPeak
|
| |
|
| Submission date |
Sep 18, 2020 |
| Last update date |
Mar 09, 2021 |
| Contact name |
Danielle R. Little |
| E-mail(s) |
Danielle.Little@stjude.org
|
| Phone |
9015953487
|
| Organization name |
St. Jude Children's Research Hospital
|
| Department |
Developmental Neurobiology
|
| Lab |
Michael Dyer
|
| Street address |
262 Danny Thomas Pl Rm D2031
|
| City |
Memphis |
| State/province |
Tennessee |
| ZIP/Postal code |
38105 |
| Country |
USA |
| |
|
| Platform ID |
GPL19057 |
| Series (2) |
| GSE158201 |
Differential chromatin binding of the lung lineage transcription factor NKX2-1 resolves opposing murine alveolar cell fates in vivo [ChIP-seq] |
| GSE158205 |
Differential chromatin binding of the lung lineage transcription factor NKX2-1 resolves opposing murine alveolar cell fates in vivo |
|
| Relations |
| BioSample |
SAMN16207216 |
| SRA |
SRX9149665 |
