GEO Accession viewer

NCBI > GEO > Accession Display

Not logged in | Login

GEO help: Mouse over screen elements for information.

Sample GSM2356809

Query DataSets for GSM2356809

Status

Public on Dec 13, 2016

Title

invitro 10U rep2, ATAC-seq

Sample type

SRA

Source name

Spleen CD8+ T cell_invitro 10U_ATAC-seq

Organism

Mus musculus

Characteristics

strain: P14 TCR Calpha deficient (C57BL/6)
cell type: Spleen CD8+ T cell
phenotype: invitro10U
infection: none

Extracted molecule

genomic DNA

Extraction protocol

ATAC-seq library preparations were performed essentially as described (Buenrostro et al., 2013), with minor modifications. Briefly, 50,000 cells were washed once in PBS before incubation for 30 minutes in lysis buffer (10 mM Tris pH 7.5, 10 mM NaCl, 3mM MgCl2, 0.1% NP-40). For one replicate of memory cells and d35 KLRG1+ cells, 10,000 cells were used. After lysis, cells were resuspended in 50 ul 1x TD Buffer containing 2.5 ul Nextera enzyme (Illumina, San Diego, CA). Transposition reactions were incubated for 30 min at 37°C before purification with QiaQuick MinElute columns (Qiagen, Valencia, CA).
Purified DNA was amplified by PCR using Kapa Hi-Fi real-time library amplification kit (Kapa, Wilmington, Massachusetts) with 10-12 cycles, according to the manufacturer’s instructions with barcoded primers, as described previously (Buenrostro et al., 2013). Amplified libraries were purified with QiaQuick MinElute columns (Qiagen, Valencia, CA) and quantitated with Kapa real-time library quantification kit (Kapa, Wilmington, Massachusetts). Paired-end sequencing was performed with the rapid run protocol with an Illumina HiSeq 2500 (Illumina, San Diego, CA) with 50 cycles in each direction.

Library strategy

OTHER

Library source

genomic

Library selection

other

Instrument model

Illumina HiSeq 2500

Description

processed data file: normCounts.tsv
SaTr-ATAC-BC6_S6

Data processing

library strategy: ATAC-seq
Basecalls and demultiplexing performed in illumina basespace
ATAC-seq reads were aligned to the mm9 genome assembly using bowtie v 1.0
Unmapped reads were trimmed of adapters and to a length of 36nt with trim_galore and remapped and merged with first bam file
bam files were filtered to extract fragments less than 100 nt, and peak summits were called using MACS2 with parameters
Peak summits from individual replicates were expanded to region of 500bp and overlapping regions from all samples were merged
Tn5 insertions sites per peak were computed for all replicates using all reads and normalized with DESeq2
Genome_build: mm9
Supplementary_files_format_and_content: tsv containing normalized counts per peak

Submission date

Oct 20, 2016

Last update date

May 15, 2019

Contact name

James Scott-Browne

E-mail(s)

[email protected]

Organization name

La Jolla Institute

Street address

9420 Athena Circle