|
| Status |
Public on Aug 08, 2013 |
| Title |
fly_Ice1_shIce1_2_ChIP_2 |
| Sample type |
SRA |
| |
|
| Source name |
S2 cell culture
|
| Organism |
Drosophila melanogaster |
| Characteristics |
cell type: S2; Schneider's Drosophila Line 2; Marco Blanchette's Lab
antibody: anti-Ice1 (CG13550, aa 1-192; Smith et al., Mol Cell. 2011 Dec 23;44(6):954-65)
|
| Treatment protocol |
Cells were fixed for 10 minutes in 1% formaldehyde, quenched with the addition of 1/10 volume of 2.5 M Glycine in PBS for 10 minutes, cells were washed twice in PBS, resuspended in hypotonic buffer and nutated for 10 minutes.3x10e7 cells were pelleted and resuspended in 1.3 ml RIPA buffer with 0.5% Sarkosyl.Cells were sonicated in polyethylene 15 ml tubes for 15 minutes, high output, in a Bioruptor Diagenode) at 4 degrees. Debris was pelleted 10 minutes at 20,000 x g 4° C. ChIP was performed with 200 µl of chromatin and 5 µg of antibodies, and scaled up 10 fold for ChIP-seq. Drosophila Ice1, Ell, and non-targeting RNAi was performed as previously described (Smith et al., Mol Cell. 2011 Dec 23;44(6):954-65) except that S2 cells were grown in SFX serum-free media from HyClone (Thermo/HyClone #SH30278.02).
|
| Growth protocol |
Drosophila chromatin immunoprecipitation was performed with S2 cells grown in Schneider's media with 10% FBS at room temperature.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP libraries were prepared with the TruSeq DNA Sample Kit (Illumina #FC-121-2001) according to Illumina's instructions.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
Chromatin IP of Ice1 after Ice1 RNAi #2
|
| Data processing |
Alignments were extended to a total fragment length of 150 bases in the orientation of the read alignment. UCSC bedGraph files were created at 25bp resolution and normalized to total alignable reads (reads-per-million).
Base-calling and quality filtering, default settings, Illumina Casava 1.8
Reads were aligned using Bowtie v0.12.7, allowing unique reads only and up to three mismatches of a 50bp seed length.
Peak detection was done using MACS, version 1.4.1 (20110622) using the indicated criteria.
RNA-seq alignments were done using TopHat v1.4.1 Differential expression analysis was done using DESeq corresponding to bioconductor version for R v2.11.1
Supplementary_files_format_and_content: http://genome.ucsc.edu/goldenPath/help/bedgraph.html
Genome_build: UCSC hg19 and UCSC dm3
|
| |
|
| Submission date |
Jun 13, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Alexander (Garrett) Garruss |
| E-mail(s) |
asg@stowers.org
|
| Organization name |
Stowers Institute for Medical Research
|
| Lab |
Shilatifard
|
| Street address |
1000 East 50th Street
|
| City |
Kansas CIty |
| State/province |
Missouri |
| ZIP/Postal code |
64110 |
| Country |
USA |
| |
|
| Platform ID |
GPL13304 |
| Series (1) |
| GSE47938 |
Multiple roles for LEC in initiation and elongation phases of snRNA gene transcription |
|
| Relations |
| BioSample |
SAMN02204117 |
| SRA |
SRX306196 |
