2024-07-22
ENA first public
ERC000011
ENA-CHECKLIST
Institute for Translational Immune-Oncology, Cancer Research Center Cologne-Essen (CCCE), University of Cologne, Cologne, Germany;  University of Cologne, Faculty of Medicine and University Hospital Cologne, Department I of Internal Medicine, Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf; Center for Molecular Medicine Cologne (CMMC), Cologne, Germany
INSDC center name
public
INSDC status
ERS20608237
SRA accession
E-MTAB-13883_3:NSG DKO 3
Submitter Id
ArrayExpress
broker name
cell_type
not collected
collection date
house mouse
common name
Protocols: Immune-reconstitution in the PBL was monitored at 8, 12, 16 and 20 post-HCT. At the endpoint analyses (week 12 or 20), PBL was collected and several tissues were biopsied (spleen, bone marrow, and thymus) and processed as previously reported (Theobald et al, 2020). The tissues were cryopreserved in cryo-medium (40% PBS; 50% Human Serum, Sigma-Aldrich, St. Louis, MO, USA; and 10% DMSO) and stored at -150 °C for further analysis. Spleen cells that were not used for FACS analysis were resuspended in 1 ml Trizol (Invitrogen, Waltham, Massachusetts, USA) and frozen at -80 °C until further use. The RNA was extracted using RNeasy MinElute kit (Qiagen, Hilden, Germany). In detail, the cells frozen in Trizol were thawed on ice and 200 µl chloroform added. The samples were centrifuged at 12,000 g for 12 min at 4 °C. Then aqueous phase was transferred to a new tube and the same volume of 70% ethanol was added. The mixture was transferred to RNeasy MinElute spin column and centrifuged at 8,000 g for 15 second at RT. The flow-through was dumped and 500 µl RPE buffer added. The samples were once again centrifuged at 8,000 g for 15 seconds at room temperature and flow-through was dumped. Then 500 µl 80% ethanol was added and the samples were centrifuge at 8,000 g for 2 min at RT. Afterwards, the samples were dried by centrifugation at max speed for 5 min. The RNA was extracted using 14 µl RNase-free water and centrifugation at max speed for 1 min. The quantity and quality of RNA was checked using photometer (NanoPhotometer N60, Implen, Munich, Germany). Library preparation and NGS analyses were carried out at the production site Cologne (Cologne Center for Genomics (CCG)).
description
not collected
geographic location (country and/or sea)
not applicable
isolate
Mus musculus
organism
Mus musculus
organism
spleen
organism part
Mus musculus
scientific_name
female
sex
NSG DKO
strain
NSG DKO 3
title