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This ChIP-seq dataset is indeed novel, and now we have included additional validation by ChIP-qPCR, which shows that low cell number ChIP maintains robust enrichment over control regions and thus is suitable for genome-wide analysis.
However, since ChIP products are special subsets of the whole genome, extensive optimization of TTAS is required for its future application on ultralow cell number ChIP products.
The mixed inputs may thus function as mutual carriers during single-tube, small cell number ChIP.
In line with the reviewer's comment we have now included a direct comparison of enrichments at control regions achieved using standard and low cell number ChIP protocol.
Increasing our stringency thresholds for SNPs in the WGS decreased the number chip loci that were present in the list of SNPs identified by sequencing (n = 329,690; ~78%), but increased concordance to ~95%.
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This method offers an alternative to other previously published methods for low cell number ChIP-seq that entails fewer cycles of amplification with associated risk of data bias.
To determine which genomic loci are marked by repressive histone methylation marks in vivo, the authors performed low cell number ChIP-seq assays for H3K27me3 and H3K9me2 in E6.25 epiblasts.
(I ) Mean C/EBPβ and AP1 complex proteins JUN and FOSL normalized (to total read number) ChIP-seq enrichments in human HepG2 and lymphoblastoid cell lines (LCLs) in a 8 kb window around the summits of ZEB1 peaks detected in LCLs.
So they went around the room to every guest picker who had pulled a numbered chip out of a cup, piecing together the original order with help from the tiny screen on a photographer's camera.
We optimized the chromatin preparation procedure to ensure maximal input material for the small-cell-number ChIP (Additional file 8: Additional Protocols and Additional file 9: Figure S6).
CEBPA-1 peaks were to a high degree shared with a biological repeat (CEBPA −2), the diluted library (CEBPA-6) and the low-cell-number ChIP (CEBPA-10K) (Additional file 6: Additional Methods).
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