{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"submitter":["Liling Xu"],"organism":["Mus musculus"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-16123"],"description":["We have employed a scRANSeq to study the difference between WT and Fip200-deficient B cell populations from mice spleen 11 days after immunization. FIP200 plays an important role in regulating LC3-dependent and LC3-independent autophage pathway, which is crucial for B cell development and differentiation. This study presents the transcriptome of Naive, GC, plasma and memory B cell populations at the peak of mice immune-response and provides new insights into the role of FIP200 in regulating plasma differentiation. FIP200f/fMb1Cre+/- (KO) mice and their littermatecontrol, FIP200f/fMb1Cre-/-(WT) were immunized with 50 ug NP29-KLH with Alum, then splenocytes isolated from WT and KO mice 11 days after immunization were enriched for GC, plasma, and memory B cells by depleting non-B cells and IgDhi B cells using magnetic beads. Those enriched samples were then barcoded and sorted for naïve, GC, plasma, and memory B cells, which were then mixed for single-cell RNA sequencing."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Sample Collection - FIP200f/fMb1Cre+/- (KO) mice and their littermatecontrol, FIP200f/fMb1Cre-/-(WT) were immunized with 50 ug NP29-KLH with Alum, then splenocytes isolated from WT and KO mice 11 days after immunization were enriched for GC, plasma, and memory B cells by depleting non-B cells and IgDhi B cells using magnetic beads. Those enriched samples were then barcoded and sorted for naïve, GC, plasma, and memory B cells, which were then mixed for single-cell RNA sequencing.","Sequencing - Libraries were sequenced via NextSeq2000 for Illumina sequencing.","Library Construction - Library was performed according to the manufactor's instruction (Chromium Single-Cell 5′ Reagent Kit v2 Chemistry, Dual Index). Briefly, the poly-A RNA form the cell lysate contained in every signle GEM was retrotranscripted to cDNA, which contains an Illumina R1 primer sequence, the 10x Barcode and Unique Molecular Identifier (UMI). The pooled barcodeed cDNA was then cleaned up with Sliane DynaBeads, amplified by PCR and the apropiated sized fragmnets were selected with SPRIselect reagent for subsequent library constrution. During the library construction, Illumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and Sample index i5 and i7 were added.","Nucleic Acid Extraction - Mixed B cells were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM)."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","Processed Data","MAGE-TAB Files"],"data_protocol":["Sequence Alignment - Sample pre-processing was performed using the Cell Ranger analysis pipeline (v7.0.0) aligning to the mm10-2020-A reference transcriptome. Sample demultiplexing, QC, normalization and clustering was performed with the R package Seurat v4.3.0 (Hao et al., 2021).","Data Transformation - ADT and HTO counts were added to the Seurat object and normalized using a centered log ratio transformation (CLR). Samples were demultiplexed using Seurat function HTODemux. Cells that expressed fewer than 300 genes, fewer than 50 housekeeping genes (list of housekeeping genes adapted from Tirosh et al., 2016) or more than 8% mitochondrial genes were removed from downstream analysis. This resulted in a Seurat object with 17731 genes across 11500 cells. Normalization was performed with sctransform (Choudhary and Satija, 2022; Hafemeister and Satija, 2019); any variance introduced by Ighv, Iglv and Igkv genes was removed by regression. Principal component analysis was performed with Seurat function RunPCA; clustering was performed with Seurat functions FindNeighbors on 40 principal components and FindClusters with the sensitivity set to 0.6. The first 40 principal components were used in Uniform Manifold Approximation and Projection (UMAP) using the Seurat RunUMAP function. Cluster markers were determined using the Seurat function FindAllMarkers."],"omics_type":["Metabolomics","Unknown","Transcriptomics","Genomics","Proteomics"],"instrument_platform":["NextSeq 2000"],"study_type":["RNA-seq of coding RNA from single cells"],"species":["Mus musculus"],"pubmed_title":["FIP200 regulates plasma B cell differentiation via mitochondrial and heme homeostasis"],"pubmed_authors":["Liling Xu1, Maria Bottermann1, Paula M. Villavicencio1, John Warner1, Stephanie R. Weldon1, Zhenfei Xie1, Andrew Filby2, Xiaotie Liu1, Ian G. Ganley3, Alison E. Ringel4, Usha R. Nair1, Facundo D. Batista1,5,6","Liling Xu"],"additional_accession":[]},"is_claimable":false,"name":"scRNASeq of Naïve, GC, plasma, memory B cells sorted from WT and Fip200-deficient mice spleen","description":"We have employed a scRANSeq to study the difference between WT and Fip200-deficient B cell populations from mice spleen 11 days after immunization. FIP200 plays an important role in regulating LC3-dependent and LC3-independent autophage pathway, which is crucial for B cell development and differentiation. This study presents the transcriptome of Naive, GC, plasma and memory B cell populations at the peak of mice immune-response and provides new insights into the role of FIP200 in regulating plasma differentiation. FIP200f/fMb1Cre+/- (KO) mice and their littermatecontrol, FIP200f/fMb1Cre-/-(WT) were immunized with 50 ug NP29-KLH with Alum, then splenocytes isolated from WT and KO mice 11 days after immunization were enriched for GC, plasma, and memory B cells by depleting non-B cells and IgDhi B cells using magnetic beads. Those enriched samples were then barcoded and sorted for naïve, GC, plasma, and memory B cells, which were then mixed for single-cell RNA sequencing.","dates":{"release":"2025-11-25T00:00:00Z","modification":"2026-05-27T15:04:25.204Z","creation":"2025-11-14T13:05:35.633Z"},"accession":"E-MTAB-16123","cross_references":{"ENA":["ERP184440"],"EFO":["EFO_0002944","EFO_0004170","EFO_0005684","EFO_0004917","EFO_0005518","EFO_0003816","EFO_0004184"]}}