cells were infected with RFP-tagged shRNA againstor control in triplicate; transduced cells were sorted 72 h post-transduction and plated in a 10 cm plate in 10% DMEM growth media. 5 days later, cells were collected for analysis by RNA-seq and ChIP-seq. >300,00 cells per replicate were collected for RNA-seq; total RNA was isolated using Quick-RNA Miniprep Kit . RNA was assessed for quality using an Agilent Tapestation; all samples had RIN > 7.
was used to process alignment files to generate ChIP-seq bed files. ChIP-seq peaks for with the parameter “-style factor” versus the appropriate ChIP input experiments as background. ChIP-seq peaks for H3K4me, H3K4me3, and H3K27ac were called using the parameter “-style histone”. database using the findMotifsGenome.pl command using default fragment size and motif length parameters. Random GC content-matched genomic regions were used as background. Enriched motifs are statistically significant motifs in input over background by aFor RNA-seq, RNA expression was quantified as raw integer counts using analyzeRepeats.pl in
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