GEO Accession viewer

NCBI > GEO > Accession Display

Not logged in | Login

GEO help: Mouse over screen elements for information.

Sample GSM2701046

Query DataSets for GSM2701046

Status

Public on Nov 22, 2017

Title

asynchronous Hi-C rep1

Sample type

SRA

Source name

late embryonic cell line

Organism

Drosophila melanogaster

Characteristics

cell line: S2R+
developmental time: embryo 20-24h
drug for synchronization: none

Growth protocol

Drosophila late embryonic S2R+ cells were seeded at the density of 1X 10^6 cells/ml and grown in Schneider's medium ( Invitrogen) with 10% heat-inactivated fetal bovine serum (BI) at 25 °C. Cells were synchronized at G1/S by incubating with 1 µM Hydroxyurea (HU) for 18 h

Extracted molecule

genomic DNA

Extraction protocol

The Hi-C chimeric DNA were prepared by "in-situ Hi-C" method with minor modifications, Briefly, nuclei released from ten million crosslinked cells were digested with DpnII (NEB). After biotin fill-in and ligation in nuclei, biotin labeled ligation product were fragmented with Cavrios M220, the molecular with size ranging from 300bp to 500bp were selected for the generation of library
The Hi-C library were prepared using NEBNext Ultra DNA Library Prep Kit for Illumina with modifications. Briefly, after end-repair and adaptor liagtion with NEB moduals, biotin labeled validated chimetric DNA were selected with Streptavidin C1 beads (Invitrogen). Hi-C library was amplified after DNA eluation by inculating at 98° C.

Library strategy

Hi-C

Library source

genomic

Library selection

other

Instrument model

HiSeq X Ten

Description

Hi-C chimeric DNA

Data processing

The two ends of the Hi-C reads were seperately mapped to the Drosophila melanogaster dm3 reference genome using Bowtie2(v2.2.9). An iterative mapping method was adopted with an initial length 25bp, if this region failed to map uniquely, an additional 25 bp was included in the next round of mapping. This procedure was continued until the full read length reached 150 bp.
Only reads with high mapping quality(MAPQ>30) were kept for further analysis. Reads mapped to the same restriction fragment and dangling reads seperated by less than 500bp and PCR duplicates were further removed.
The Hi-C contact maps were normalized using ICE(Imakaev et al, 2012), restriction fragments were treated as units instead of equally spaced bins.
Domains were called using Armatus(Filippova et al, 2014) with scaling parameter, gamma, set to 0.9. Domains interrupted by gaps were modified locally using gamma 0.6.
Adapters and low quality reads were removed by cutadapt (v1.8.3) and Trimmomatic (v0.36) using default parameters. Valid paired reads were mapped to the Drosophila melanogaster reference genome dm3 and Flybase gene annotation v5.57 using Tophat2 (v2.0.9) with default options and transcripts were assembled using Cufflinks (v2.2.1). Two replicates were merged for further analysis.
Genome_build: dm3
Supplementary_files_format_and_content: S2Rplus_G1S_domain.txt is a tab-delimited text file contains domain positions in synchronized G1S S2R+ cells. G1S_genes.txt is a tab-delimited text file contains positions, ids, names and expression levels(FPKM) of Flybase r5.57 annotated genes .

Submission date

Jul 12, 2017

Last update date

May 15, 2019

Contact name

Qi Wang

E-mail(s)

rubbyqi@sjtu.edu.cn

Organization name

Shanghai Jiaotong University

Street address

800 Dong Chuan Rd.