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| Status |
Public on Dec 23, 2015 |
| Title |
ATAC-seq Acinar2 |
| Sample type |
SRA |
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| Source name |
pancreatic islets
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| Organism |
Homo sapiens |
| Characteristics |
cell type: acinar
donor: ACFV445
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| Treatment protocol |
Islets underwent fluorescence-activated cell sorting using cell-surface antibodies (HIC1-2B4, HIC3-2D12, and HIC1-1C10) to separate alpha, beta, and acinar cells.
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| Growth protocol |
Islets were cultured either in CMRL1066 with 0.5% human albumin, 2 mM L-glutamine, 1% heparin, and 1% penicillin/streptomycin or in Prodo PIM(S) media supplemented with 5% PIM(ABS) and 1% PIM(G) at 37oC with 5% CO2.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Nuclei were isolated from cells by lysing in 50 ul/50,000 cells of lysis buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% NP-40), followed by centrifugation at 500 x g for 10 minutes at 4 degrees Celsius. Transposition was performed using 50 ul/50,000 cells of transposition reaction mix (25 ul TD buffer, 2.5 ul Tn5 transposase, 22.5 ul nuclease-free water) from the Illumina-Compatible Nextera DNA Sample Prep Kit (Part# FC-121-1030) for 30 minutes at 37 degrees Celsius. DNA was then purified using the Qiagen MinElute PCR Purification Kit (Part# 28004) and eluted in 10 ul of elution buffer.
Libraries were prepared using Nextera barcoded primers and NEBNext High-Fidelity 2X PCR Master Mix (Part# M0541).
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Library strategy: ATAC-seq
ATACseq: Illumina adapters were trimmed by Trimmomatic, and the 50 bp reads were trimmed by 10 bp with fastx_trimmer, then all reads were combined for each sample and aligned to hg18 with bowtie2 using default settings. Reads mapped to mitochrondrial DNA and low quality reads (MAPQ<10, duplicates and reads in Encode blacklist regions) were removed. All reads left were offset by +4 bp for + strand and −5 bp for – strand. Peaks were called for each sample using MACS2 with parameters “-q 0.05 --nomodel --shift 37 --extsize 73”.
RNAseq: High quality RNA-seq reads were aligned to the human genome and transcriptome (hg18) using the RNA-seq Unified Mapper (RUM). Aligned reads were filtered for rRNA reads using BLAST alignment against the known rRNA sequences, and all reads aligning to the mitochondrial genome were also discarded. Aligned reads were used to determine unambiguous read counts for each gene in the refGene database using HTSeq, i.e., only reads uniquely assigned to a single gene were counted. All differential expression analysis was performed using the GLM functions in the EdgeR package.
Genome_build: hg18
Supplementary_files_format_and_content: bed files from MACS2 for ATACseq peak summit locations; tab-delimited text files include read counts for each RNAseq ample
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| Submission date |
Dec 22, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Paul Wang |
| E-mail(s) |
zhipwang@mail.med.upenn.edu
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| Phone |
215573117
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| Organization name |
University of Pennsylvania
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| Street address |
3700 Hamilton Walk
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| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19104 |
| Country |
USA |
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| Platform ID |
GPL18573 |
| Series (1) |
| GSE76268 |
Integration of ATAC-seq and RNA-seq Identifies Human Alpha Cell and Beta Cell Signature Genes |
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| Relations |
| BioSample |
SAMN04362228 |
| SRA |
SRX1497366 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1978249_ACFV445acinar2_summits.bed.gz |
731.2 Kb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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