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Sample GSM2598433 Query DataSets for GSM2598433
Status Public on May 10, 2017
Title CTCF.sample2.unbound
Sample type SRA
 
Source name In vitro protein-DNA binding reactions
Organism Mus musculus
Characteristics dna library composition: Compared to CTCF sample1, the barcodes labeling unmethylated and M.sssI treated dsDNA were swapped to each other.
recombinant protein used: mouse CTCF (Finger 1 to Finger 9)
Treatment protocol All EMSA reactions were done in 9% Tris-glycine gel, 200V, 30mins
Extracted molecule genomic DNA
Extraction protocol After EMSA, DNA fragments were cut and extracted by gel extraction buffer at 50C for 30mins.
For each portion of DNA fragment, PE1 and PE2 PCR primers were used to PCR amplify extracted DNA by at least 8 cycles (95C 15s, 52C 15s, 72C 30s)
For each particular sample, there are two raw data files, i.e., bound reads and unbound reads with different Illumina Index. The processed data file was based on analysis of Bound and Unbound data together.
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina MiSeq
 
Description CTCF_sample2_processed
Data processing Library strategy: Methyl-Spec-seq
Methylation property identification: For any particular DNA variant inside library, it carries particular barcode to indicate its methylation property, i.e., unmethylated, top hemimethylated, bottom hemimethylated, duplex methylated. Thus acording to the unique methylation barcode information, each read was identified and sorted to its methylation type.
Variant reads counting and sorting: For each randomized binding site, its number in Bound and Unbound portions were counted separately. The Bound/Unbound ratio was supposedly porportional to its relative binding affinity, and the negative logarithm of this ratio indicates the relative binding energy in kT unit.
Energy logo visualization: Based on the binding energy for particular reference and all its single variants, an energy logo can be generated to indicate the specificity of particular protein-DNA interactions. Futhermore, besides conventional G-T-A-C bases, we used two extra “M" and ”W" to represent methylated cytosine on upper and lower strand of DNA respectively, in which the height of M or W indicate the extra binding energy contribution between methylated and unmethylated base.
Supplementary_files_format_and_content: All processed data were saved in .excel files, including the methylation property, bound reads, unbound reads, Bound/Unbound ratio, relative binding energy for each variant.
 
Submission date May 04, 2017
Last update date May 15, 2019
Contact name Zheng Zuo
E-mail(s) zeropin@live.cn
Organization name Stanford university
Department Genetics
Street address Stanford
City Palo Alto
State/province CA
ZIP/Postal code 94040
Country USA
 
Platform ID GPL16417
Series (1)
GSE98542 Methyl-Spec-seq control data sets
Relations
BioSample SAMN06890678
SRA SRX2783745

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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