2024-06-15
ENA first public
ERC000011
ENA-CHECKLIST
Institute of Innate Immunity, Medical Faculty, University of Bonn
INSDC center name
public
INSDC status
ERS20164468
SRA accession
E-MTAB-14126:Anti-GPIb_PBS_N2_R1
Submitter Id
12
week
age
ArrayExpress
broker name
FACS-Sorted Ly6G-I-A/I-ECD45+  CD11b+CD115+ monocytes
cell_type
not collected
collection date
house mouse
common name
Protocols: In vivo, platelet depletion was performed in female C57BL/6J mice (12 weeks) by intravenous (i.v.) injection of 2 mg/kg of polyclonal rat anti-mouse GPIbα (R300) or same amounts of a polyclonal rat IgG none-immune (C301) antibody as control (Emfret Analytics). Mice were then challenged 12 hours later with i.v. injection of 2 mg/kg LPS or PBS as control. Blood was collected after 3 h, and PBMCs were isolated by density gradient centrifugation in Ficoll Paque PLUS. Mouse monocytes were separated from PBMCs by FACS-sorting gating on Ly6G−, IA/IE−, CD45+, CD11b+, and CD115+ cells and sorted directly into QIAzol lysis reagent. Lysates were frozen in liquid nitrogen and sent to GENEWIZ Azenta Life Sciences for bulk RNA sequencing.   For bulk RNA sequencing, samples were processed according to GENEWIZ (Azenta Life Sciences) using TRIzol extraction protocol. QC was done on QuBit and TapeStation.   RNA extraction was performed, followed by a library preparation (Azenta, GeneWiz).  SmartSeq HT library preparation kit with maximum input volume as the RNA amount was below QuBit detection limit.  mRNA sequencing via polyA selection: 1) mRNA Enrichment, mRNA Fragmentation, and Randon Priming; 2) First and second strand cDNA synthesis; 3) End Repair, 5' Phosphorylation, and dA-Tailing; 4) Adaptor Ligation, PCR Enrihment, and Sequencing.
description
adult
developmental stage
wild type genotype
genotype
not collected
geographic location (country and/or sea)
not applicable
isolate
Mus musculus
organism
Mus musculus
organism
blood
organism part
3
replicate
Mus musculus
scientific_name
female
sex
PBS
stimulus
C57BL/6J
strain
Anti-GPIb_PBS_N2_R1
title
Anti-GPIb
treatment