|
| Status |
Public on Mar 18, 2016 |
| Title |
Hi-C library HS1 |
| Sample type |
SRA |
| |
|
| Source name |
female embryonic cells
|
| Organism |
Drosophila melanogaster |
| Characteristics |
cell type: embryonic (established from 6-12hours embryo)
cell line: Kc167
hi-c enzyme: HindIII
treatment: Heat shocked
|
| Treatment protocol |
One hundred million Kc167 cells were suspended in 20ml of CM3 medium at room temperature mixed with equal volume of 48 degree medium and incubated in 37 degree water bath for 20 minutes. Heat shocked cells were then fixed by adding 0.6 volume of 4 degree medium contain 2.7% formaldehyde to a final concentration of 1%, and incubated at room temperature for 10 min. The reaction was stopped by adding 1/10 volumes of 1.25 M glycine solution and incubated for 5 min before centrifugation at 3000 rpm for 10 min.
|
| Growth protocol |
Fruit fly embryonic Kc167 cells were grown to confluence in CCM3 serum-free insect medium (HyClone SH30065.01) at 25oC and collected for Hi-C experiments.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Hi-C experiments were carried out as described (Lieberman-Aiden et al., 2009) with some modifications. Cells were lysed then the nuclei were treated with 0.1% SDS at 65 degree for 10mins and the SDS was quenched by adding Triton X-100 to final 1% concentration. Nuclei was digested in 1XNEB (New England Biolaboratory) with 400U of HindIII. Digested DNA was blunted with dATP, dGTP, dTTP and biotin labeled dCTP. DNA wan then ligated and puried. biotin-labled ligated DNA was purified with streptavidin beads. Illumina paired-end adaptor linkers were ligated to DNA pulled-down and amplified and size selected for sequencing.
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~300 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the HiSeq2000 following the manufacturer's protocols. Hi-C DNA was ligated with Illumina adaptors and then amplified, purified. ChIP DNAs were ligated with six different Illumina index adaptors and amplified, purified.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
Heat shocked
supplementary file: AllHS_hotint_NB_10000rs_allchr.txt
|
| Data processing |
Library strategy: Hi-C+
Hi-C datasets were normalized as described (Yaffe and Tanay, Nature Genetics, 2011)
base-calling was carried out by HudsonAlpha Genomic Service Center
Paired reads were aligned to the Drosophila reference genome (Dm3) using Bowtie 0.12.7 (Langmead et al., 2009) and filtered to remove any pair with one or two ends aligned to multiple locations.
Genome_build: Dm3
Supplementary_files_format_and_content: Six ChIP-seq wig files of protein binding peaks called by MACS, one ChIP input control. One excel file contains listes of long-range interactions identified before and after heat shock at 10kb resolution.
Supplementary_files_format_and_content: AllHS_hotint_NB_10000rs_allchr.txt: 10kb resolution interactions for heat shocked Hi-C in Kc167 cells
Supplementary_files_format_and_content: All3_hotint_NB_10000rs_allchr.txt: 10kb resolution interactions for normal Hi-C in Kc167 cells
Supplementary_files_format_and_content: hsBEAF_treat.wig: BEAF peaks in heat shocked Kc167 cells
Supplementary_files_format_and_content: hsCP190_treat.wig: CP190 peaks in heat shocked Kc167 cells
Supplementary_files_format_and_content: hsCTCF_treat.wig: CTCF peaks in heat shocked Kc167 cells
Supplementary_files_format_and_content: hsPolII_treat.wig: RNAPII peaks in heat shocked Kc167 cells
Supplementary_files_format_and_content: hsSuHw_treat.wig: SuHw peaks in heat shocked Kc167 cells
Supplementary_files_format_and_content: input.wig: input DNA
Supplementary_files_format_and_content: polII_treat.wig: RNAPII in normal Kc167 cells
|
| |
|
| Submission date |
Mar 19, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Chun-hui Hou |
| Organization name |
Emory University
|
| Street address |
1510 Clifton Rd NE
|
| City |
Atlanta |
| ZIP/Postal code |
30322 |
| Country |
USA |
| |
|
| Platform ID |
GPL13304 |
| Series (1) |
| GSE45293 |
Temperature Sensitive Remodeling of Fruit Fly Physical Chromosomal Domains upon Heat Shock |
|
| Relations |
| SRA |
SRX252222 |
| BioSample |
SAMN01984531 |
