|
| Status |
Public on Jul 20, 2013 |
| Title |
nlsSwi6KR25A_H3K9me2_ChIPSEQ_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
nlsSwi6KR25A H3K9me2 ChIPSEQ
|
| Organism |
Schizosaccharomyces pombe |
| Characteristics |
genotype: nlsswi6KR25A
chip antibody: H3K9me2
chip antibody manufacturer: WAKO
chip antibody catalog #: MABI0307
chip antibody lot #: 302-32369
|
| Growth protocol |
S.pombe strains were grown in YES media. ~150x10^7 cells were harvested from o/n cultures and processed for ChIP
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated S. pombe cells and histone-DNA complexes were isolated with antibody.
ChIP-seq libraries were generated using an Illumina-based protocol using custom reagents and barcoded adapters. Briefly, immunoprecipitated 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 fragments were size selected to remove excess adapters and adapter modified DNA fragments were enriched by PCR amplification with Illumina primers for 18 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified ChIP-seq libraries thus generated were validated and subsequently captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Illumina HiSeq 2000 system following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
CCACTCT
|
| Data processing |
All samples were barcoded at the 3'end of the 5'adapter using a hamming distance two code with a 3' CT (GTGCGCT, GCCACCT, AGAAGCT, ATCTTCT, TGCCACT, CACGGCT, CGGTCCT, CCACTCT) and sequenced (with other samples) in two lanes of an HiSeq 2000 instrument. Individual reads were assigned to their sample based on the first 7 nucleotides containing the barcode. The 3' adapter was removed by aligning it to the read allowing one or two mismatches in prefix alignments of at least 7 or 10 bases respectively. Low complexity reads were filtered out based on their dinucleotide entropy (removing <1% of the reads). All the reads that were shorter than 14 nucleotides were removed.
Genome_build: Initial identification of elements is based on the features described in the GFF file “pombe_09052011.gff” (downloaded from ftp://ftp.sanger.ac.uk/pub/yeast/pombe/GFF).
Supplementary_files_format_and_content: wig file for multiple tracks
|
| |
|
| Submission date |
Dec 11, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Tim Roloff |
| E-mail(s) |
tim.roloff@fmi.ch
|
| Organization name |
FMI
|
| Department |
Functional Genomics
|
| Street address |
Maulbeerstrasse 66
|
| City |
Basel |
| ZIP/Postal code |
4058 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL13988 |
| Series (2) |
| GSE42848 |
Genome wide map of heterochromatin state in fission yeast Schizosaccharomyces pombe |
| GSE42850 |
Heterochromatin state in fission yeast Schizosaccharomyces pombe |
|
| Relations |
| SRA |
SRX209514 |
| BioSample |
SAMN01830784 |
