NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM301565 Query DataSets for GSM301565
Status Public on Jul 03, 2008
Title HEK Input DNA rep3
Sample type SRA
 
Source name Input DNA
Organism Homo sapiens
Characteristics n/a
Growth protocol HEK 293T cells were cultured in parallel in 2*150 cm3 flasks with DMEM (penicillin-streptomycin) (Gibco) supplemented with 10% Fetal Calf Serum (FCS) (Biochrom) and Glutamax (1x) (Gibco). In short, 5-10 ? 107 293T/17 cells were cross-linked with 1% formaldehyde for 10 min and the reaction stopped by adding 1/20 volume of 2.5 M glycine. The cross-linked material was washed with PBS, lysed, and sonicated to an average size of 300 bp.
Extracted molecule genomic DNA
Extraction protocol The sheared chromatin was incubated with 10 µg specific antibody to RNA polymerase II (ab5408, recognizing the hypophosphorylated form of RNA pol II) coupled to protein G magnetic beads (Invitrogen) for 16 h at 4°C. An aliquot of the input DNA was saved prior to immunoprecipitation as reference sample. The DNA was finally recovered, after washing (wash buffer: 50 mM Hepes-KOH, pH 7.6), 500 mM LiCl;1mM EDTA; 1% NP-40; 0.7% Na-Deoxycholate) and elution, by reversing the cross-linking overnight at 65° C in the elution buffer (50 mM Tris-HCl, pH 8.0; 10mM EDTA; 1% SDS). 80 ng of input DNA and 10 ng of ChIP DNA (see above) were taken for library preparation. The amplification was performed prior to gel size fractionation. 18 cycles of PCR with Illumina PCR primers were performed using an elongation time of 45 seconds. PCR-products were size fractionated and 150-300bp fragments were excised.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina Genome Analyzer
 
Description raw sequence 070929_6
Data processing After mapping of the reads to the genome, the replicates were pooled and resulted in 3,198,755 unique reads for two lanes of the RNA Pol II experiment and in 8,017,459 unique reads for three lanes of the control input DNA. The reads were sorted by their position on the chromosome and the start positions were used for the analysis. The RNA-Seq reads were analyzed for clusters of 10 reads in an initial window of 100 base pairs. 198 of the Pol-II clusters were overlapping with one of the input control clusters and thus were discarded.
 
Submission date Jun 30, 2008
Last update date May 15, 2019
Contact name Marc Sultan
E-mail(s) sultan@molgen.mpg.de
Organization name Max Planck Institute for molecular genetics
Street address Ihnestr. 73
City Berlin
ZIP/Postal code 14195
Country Germany
 
Platform ID GPL9052
Series (1)
GSE11892 A Global View of Gene Activity and Alternative Splicing by Deep Sequencing of the Human Transcriptome
Relations
SRA SRX003881
BioSample SAMN02195258

Data table header descriptions
SEQUENCE dots in sequences denote ambiguous base calls
COUNT number of times sequenced

Data table
SEQUENCE COUNT
..CGTACATCCAGTCTGTGATATTGTTCTTAATATT 0
..GAAATGTTGATTATTTTACTGGCACATCTTTGGC 1
..TAGCATTCAGGTTTGCGGGATGGGGCCTCTCACT 0
..TCACGATTACCCCGCTCAACGCCCAAACCCAAAC 1
..TTCTTGGTGTGGGGCTCATGCACTTATTTATTCC 1
.A.GTAAGGAGCTGTCTTGTGGGGTGAGACATTGAT 1
.A.TAAAAAAGTTTTGTCACTTAACAAGATAAACCC 1
.A.TAATTTTCATTTACTAATAGAACAAATATCTAA 1
.AA.AAGTTTCTGGGGTTTACCTCTCCGTGCTACTC 1
.AA.ATTTCATTGGTAGTGTTTTAGGACAATATCTT 1
.AA.CACAACGCCATTGAGGATTAAAGGCTGACACT 1
.AA.CACATAAGCATTTCCTTTAGGCTCTGGCTTCT 1
.AA.CCTATTCCCATGTCGTTGCGATACCCCCCCAT 1
.AA.GAGTGGAATTGGATAGTTTTATAACACAAAGA 1
.AA.GAGTGTAGTGGCGTGATCTCTGCCCAACGCCA 1
.AA.GCCAGGATAACCCGGTGGTCAAACTTTCTGCA 1
.AA.GGCACAACTGAAGCACACGCCCCTCCCCCCCC 1
.AA.GGGCAGCATAAGTAAACCTGATCTGCCAGGAT 1
.AA.TTTTTAACTAAGGGTGATGGACCTCTTGGGCT 1
.AAAAAAAAAATTGCAATTTGGACTCTTGTGGAGAA 0

Total number of rows: 12814371

Table truncated, full table size 449565 Kbytes.




Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data included within Sample table

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap