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Sample GSM4523851 Query DataSets for GSM4523851
Status Public on Feb 28, 2021
Title healthy control NeuN-positive Rep3
Sample type SRA
 
Source name cortical neuronal nuclei
Organism Homo sapiens
Characteristics disorder: healthy control
tissue: NeuN-positive
Treatment protocol Isolation of nuclei was performed as previously described (Krishnaswami et al., 2016) with minor modifications. Briefly, 50-250mg of pre-frontal cortical grey matter was homogenised in homogenisation buffer (250mM sucrose, 25mM KCl, 5mM MgCl2, 10mM Tris buffer pH 8.0, 1μM DTT, 1X Proteinase Inhibitor w/o EDTA (Roche), 0.4U μl-1 RNaseIn (ThermoFisher) 0.2U μl-1 Superasin (ThermoFisher), 1μM DAPI) and centrifuged through an iodixanol gradient (Sigma). Pelleted nuclei were washed and then stained with NeuN antibody (Abcam, ab190195) in staining buffer (PBS, 1%BSA, 0.2U/μl RNaseIn (Thermofisher), NeuN antibody (1:200)) for one hour at 4oC. For negative controls antibody was excluded. Nuclei were then sorted on BD Fusion, and collected in wash buffer (PBS, 1%BSA, 0.2U/μl RNaseIn (Thermofisher)), before proceeding to downstream RNA extraction.
Extracted molecule total RNA
Extraction protocol RNA was purified using the PicoPure RNA isolation kit (ThermoFisher) following the manufacturer's instructions.
40ng of total RNA was used to prepare libraries using the NEBNext® Ultra™ II Directional RNA Library Prep Kit for Illumina (ribozero ribosomal RNA depletion), following the manufacturer recommendations. Library quality and quantity were assessed on a Bionalayser and Qubit respectively. Libraries were sequenced on an Illumina Hiseq2500 (v4 chemistry) and at least 40million paired end 100bp reads per sample were generated per library..
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2500
 
Description RNA-seq of healthy control cortical neuronal nuceli
Ctrl_NueNngeVSNueNpos_GeneCounts.csv
1499NeuNpos
Data processing Illumina CASAVA 1.8.4 software used for basecalling.
Reads were mapped to hg38 whole genome using tophat2 (2.0.11) with argument “--library-type fr-firststrand"
Gene annotations were obtained from iGenome UCSC hg38. Gene-based read count were obtained using featureCounts function from Rsubread Bioconductor package 
Genome_build: hg38
Supplementary_files_format_and_content: csv file includes Gene counts for each Sample
 
Submission date May 08, 2020
Last update date Feb 28, 2021
Contact name LMS Bioinformatics Core
E-mail(s) bioinformatics@lms.mrc.ac.uk
Organization name MRC London Institute of Medical Sciences
Department Bioinformatics Core
Street address Hammersmith Hospital Campus, Du Cane Road
City London
ZIP/Postal code W12 0NN
Country United Kingdom
 
Platform ID GPL16791
Series (2)
GSE150121 Human cortical nuclei RNAseq
GSE150130 Partial rescue of neuronal genes deregulated in CdLS by cohesin
Relations
BioSample SAMN14854893
SRA SRX8293771

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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