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RNASeq of RNA from naïve B cells and T cell blasts from individuals with AIOLOS N160S mutation and healthy controls

ABSTRACT: RNA-seq was performed using RNA extracted from enriched T cell blasts (CD3 T cells were enriched from the T-cell blasts, purity >90%, Stemcell Technologies, 19051) or naïve B cells (purity >90%, Stemcell technologies, 19254) for the following study groups: healthy controls (HC, n=3-4) and patients with AIOLOS N160S (n=1 for naïve B cells [A.III.1] and n=2 for T cell blasts [A.II.2 and A.III.1]). Libraries were prepared using the AmpliSeq for Illumina Transcriptome Human Gene Expression panel, which captures 20,802 genes (>95% of human RefSeq genes). RNA-seq was performed on the Illumina HiSeq 2500 (Illumina; AmpliSeq for Illumina/HiSeq 2500). Demultiplexed reads were mapped to the hg19 genome using the splice-aware aligner Tophat (Trapnell et al., 2009). Gene-level counts data were generated using the Rsubread feature counts a read summarization program that counts mapped reads for genomic features such as genes(Liao et al., 2019). Differential expression analysis was performed using R (v.3.5.3) and DESeq2 (v.1.22.2)(Love et al., 2014).

ORGANISM(S): Homo sapiens

PROVIDER: GSE183966 | GEO | 2021/09/14

REPOSITORIES: GEO

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Quantitative analysis of chromatin occupancy (ChIP-Seq) from wild-type mice and Kdm5c KO mice in standard conditions (H3K4me3, H3K4me1, and Kdm5C) or upon novelty exposure (H3K4me3), and control mice and Kdm5c ifKO mice in basal state (H3K4me3).

Project description:Chromatin immunoprecipitation (ChIP) experiments were conducted as previously described (Ito et al, 2013) using anti-H3K4me3 (Millipore, #07-473), anti-H3K4me1 (Abcam, #ab8895), or anti-Kdm5C (Iwase et al., 2016). Hippocampi derived from two different animals were pooled together for each sample and two independent biological replicates per condition were sequenced according to manufacturer instructions in a HiSeq2500 apparatus (Illumina, Inc). Information on library preparation method, size of the libraries, and mapping to reference genome can be found in Supplementary Material accompanying the manuscript. ChIP-seq reads were aligned to the mouse genome (Mus_musculus.GRCm.38.83) using bowtie2 (v2.2.9) (Langmead and Salzberg, 2012) and further processed using samtools (v1.3.1) (Li et al., 2009). Peak calling was performed using MACS2 (v2.1.0) (Zhang et al., 2008) with default parameters except for Kdm5c that were as follows: -q 0.01 --nomodel --extsize 131 --broad --broad-cutoff 0.1. Read counts on aligned bam files were performed using Rsubread (v1.22.3) (Liao et al., 2014). Differential peak methylation analysis for H3K4me3 chromatin mark was performed using DESeq2 (v1.10.0) (Love et al., 2014) of the bioconductor suite (Huber et al., 2015) in the R (v3.3) statistical computing platform. For consideration of differentially methylated regions between conditions, we used adjusted p-value < 0.05 as indicated in the manuscript.

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| PRJEB7907 | ENA

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