GSE106211
ChIP-seq targeting Myc-tagged UL34 from human fibroblasts infected with human cytomegalovirus at 48 hours post infection
Purpose: To identify interactions of the viral DNA-binding protein, UL34, with the viral genome during lytic-phase replication. Methods: Human fibroblasts were infected with human cytomegalovirus expressing Myc-tagged UL34 in biological duplicate.  At 48 hours post infection, ChIP was performed with anti-Myc tag antibody (Cell Signaling) and the resulting DNA fragments were sequenced on Illumina MiSeq.  Reads were trimmed and duplicates removed.  Reads passing quality filter were aligned to a custom reference genome containing both the human genome with the human cytomegalovirus genome as an additional chromosome to prevent read assignment bias with CLC Genomics Workbench verison 9.0.  Enrichment of UL34 in the viral origin of lytic replication (oriLyt) was calculated by dividing read counts from the ChIP''d samples (Myc-UL34-1, Myc-UL34-2) by those from non-ChIP whole cell lysate (Input) with a 125bp sliding window (25bp overhang) after adjusting for total read count variances.  An additional sample containing DNA that was subjected to the ChIP protocol in the absence of antibody (No Antibody) was compared to the Input to determine background enrichment. Results: Several peaks were identified when comparing the Myc-UL34-ChIP samples to Input.  UL34 is bound to the viral oriLyt at 48 hours post infection near the three previously defined UL34-binding sites to varying degrees, as well as additional locations within the region.  No peaks were identified when comparing the No Antibody and Input samples. Overall design: Occupancy of HCMV UL34 in the viral oriLyt.