B cell transcriptomic analysis upon YY1 deletion

Mu-IgM/anti-CD40 stimulated follicular B cells were isolated from 3 month old female or male yy1^flox/flox animals (three separate animals each) treated with or without TAT-CRE. About 48 hours post-stimulation and YY1 deletion, total RNA was isolated using TRIzol (Invitrogen) and libraries were prepared with an Illumina TruSeq Stranded mRNA LT kit. Samples were run on Illumina NextSeq 500 and bioinformatic analyses were conducted as previously described [75 (link)]. Briefly, R (v3.3.1), RStudio (v1.0.44), and the Bioconductor suite of packages (http://www.bioconductor.org) were used to perform X-linked gene expression analyses. RNA-Seq reads were aligned with Tophat to the GRCm38/mm10 mouse reference genome and HTSeq-count and edgeR were used to normalize data. Statistical significance was determined with log2 fold change (lfc) and false discovery rates (FDR) (FDR <0.05, lfc > 0.5 or <-0.5). Heatmaps were constructed in RStudio (gplots, RColorBrewer), and expression levels for differentially expressed X-linked genes are shown. Gene Ontology (GO) term enrichment was performed using DAVID v6.8. A selection of Functional Annotation Clustering terms with an enrichment score >1 (p < 0.05) are shown. The RNAseq data are available in the Gene Expression Omnibus (GEO) database under the accession number GSE104097.