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SRX20301954: GSM7328890: Rat CD4+ T-MBP cells with validation library, blood, rep R3; Rattus norvegicus; OTHER
1 ILLUMINA (Illumina HiSeq 1500) run: 16.1M spots, 803M bases, 321.9Mb downloads

External Id: GSM7328890_r1
Submitted by: Institute for Clinical Neuroimmunology, Biomedical Center, University Hospital of the Ludwig-Maximilians University Munich
Study: Genome wide (GW) and validation CRISPR screens on encephalitogenic CD4+ T cell migration into the CNS
show Abstracthide Abstract
To identify genes driving encephalitogenic CD4+ T cell migration into the CNS, we performed a genome-wide CRISPR screen and a subsequent validation screen For the genome-wide CRISPR screen, up to 4 sgRNA per gene and 800 non targeted controls were included, for a total of 87690 individual sgRNAs, and for the validation screen, up to 6 sgRNA per gene and 241 non targeted controls for a total of 12000 individual sgRNAs Overall design: Comparison of sgRNA distribution by pairwise comparisons across blood, spleen, spinal cord leptomeninges and spinal cord parenchyma
Sample: Rat CD4+ T-MBP cells with validation library, blood, rep R3
SAMN35052128 • SRS17627810 • All experiments • All runs
Library:
Name: GSM7328890
Instrument: Illumina HiSeq 1500
Strategy: OTHER
Source: GENOMIC
Selection: other
Layout: SINGLE
Construction protocol: Blood was drawn by heart puncture into a heparinized syringe. Lymphocytes were isolated from blood by a Nycoprep gradient. First, the blood was diluted with an equal volume of PBS and overlaid onto Nycoprep. After centrifugation at 800 g, room temperature for 30 min with mild acceleration and brake, lymphocytes were collected from the interface. BFP+ T cells were FACS sorted. Genomic DNA (gDNA) from lymphocytes or sorted TMBP cells was isolated with the DNeasy Blood and Tissue Kit (Qiagen). A one-step PCR amplification was performed with Q5 High Fidelity DNA Polymerase by using 2.5µg of gDNA per reaction with Fwd-Lib (mix of 8 staggered primers) and Rev-Lib (consists of 8bp of unique barcode) primers for a total of 24 cycles. Illumina adapters were introduced together with the amplification primers. All primer sequences are listed in Supplementary Table 1. The amplified DNA amplicons were purified with SPRIselect (Beckman Coulter) with a ratio of 1:0.8 (DNA to beads) and eluted in nuclease-free water. The presence of ~250bp DNA amplicons was confirmed and the concentration was measured with Agilent Bioanalyzer on DNA 1000 Chips (5067-1504).
Runs: 1 run, 16.1M spots, 803M bases, 321.9Mb
Run# of Spots# of BasesSizePublished
SRR2451765516,060,441803M321.9Mb2023-08-01

ID:
27723126

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