Stage 4–6 embryos were collected at 25°C, and embryo stage was confirmed microscopically. Embryos were fixed in 1.8% formaldehyde for 15 min at room temperature (details are in the Supplemental Material ). ChIP was performed as described previously (Huang et al. 2014 (link)).
The antibodies used for ChIP included 10 µg of rabbit α-Sgf11, 20 µg of rabbit α-Ada2b, 20 µg of rabbit α-Spt3, 20 µg of rabbit α-Non-stop (details are in theSupplemental Material ), 3 µg of mouse α-Pol II (4H8) (Abcam, ab5408), 5 µg of mouse α-ubH2B (EMD Millipore, 17-650), 5 µg of rabbit α-H2B (Abcam, ab1790), 5 µg of rabbit α-H3K9ac (Abcam, ab4441), 5 µg of rabbit α-H3K14ac (Abcam, ab52946), and 5 µg of rabbit α-H3 (Abcam, ab1791).
Reads of 51 bases from an Illumina HiSeq 2500 were aligned to version dm3 of the Drosophila genome from University of California at Santa Cruz using Bowtie using parameters --best --strata -k 1 -m 3. The resulting BAM files were analyzed in R using Bioconductor to generate coverage and normalize the data in reads per million. Locations of enrichment for each protein were identified using MACS2 with default parameters.
ChIP-seq and RNA-seq data are available in Gene Expression Omnibus under accession number GSE98865.
The antibodies used for ChIP included 10 µg of rabbit α-Sgf11, 20 µg of rabbit α-Ada2b, 20 µg of rabbit α-Spt3, 20 µg of rabbit α-Non-stop (details are in the
Reads of 51 bases from an Illumina HiSeq 2500 were aligned to version dm3 of the Drosophila genome from University of California at Santa Cruz using Bowtie using parameters --best --strata -k 1 -m 3. The resulting BAM files were analyzed in R using Bioconductor to generate coverage and normalize the data in reads per million. Locations of enrichment for each protein were identified using MACS2 with default parameters.
ChIP-seq and RNA-seq data are available in Gene Expression Omnibus under accession number GSE98865.
