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| Status |
Public on Jun 25, 2019 |
| Title |
4504_Schizo_NeuN |
| Sample type |
SRA |
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| Source name |
frozen brain tissue_Schizo_NeuN
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| Organism |
Homo sapiens |
| Characteristics |
subject id: 4504
cell type: NeuN
diagnosis: Schizophrenia
Sex: M
age: 55
pmi: 10.7
hemisphere: L
brodman area: Brodmann Area 46
brainbank: HSB
rin: 5.2
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| Extracted molecule |
total RNA |
| Extraction protocol |
nuclear extraction: Tissue isolated from Brodmann Area 46 from 40 individuals with schizophrenia and 49 matched controls.700 mg of frozen postmortem brain was homogenized with lysis buffer using a dounce homogenizer. Brain lysate was placed on a sucrose solution to create a concentration gradient. Ultracentrifuge was used to separte nuclei and cytoplasmic fraction.
FANS protocol: Nuclei were resuspended in PBS plus RNAse inhibitors and incubated with mouse alexa488 conjugated anti-NeuN (1:200) (#MAB377X, Millipore, Billerica, MA) and rabbit alexa555 conjugated anti-OLIG2 (1:75) (#AB9610-AF555, Millipore) antibodies with 0.5% BSA. Immuno-labeled nuclei were collected as NeuN-positive or OLIG2-positive populations by fluorescence-activated cell sorting (FACS).
RNA extraction: Total RNA were purified from each nuclei population using a ZR-Duet DNA/RNA MiniPrep (Plus) kit according to the manufacturer's instruction. 200 ng total RNA from each sample was treated for ribosomal RNA (rRNA) removal using the Low Input RiboMinus Eukaryote System v2 (#A15027, ThermoFisher) according to the manufacturer's instruction.
Total RNAs with an average RIN value of 7.5±0.16 were used for RNA-seq library preparation. 50 ng of total RNA after rRNA removal was subjected to fragmentation, first and second strand syntheses, and clean up by EpiNext beads (#P1063, EpiGentek, Farmingdale, NY). Second strand cDNA was adenylated, ligated and cleaned up twice by EpiNext beads. cDNA libraries were amplified by PCR, and cleaned up twice by EpiNext beads. cDNA library quality was quantified by a 2100 Bioanalyzer using an Agilent High Sensitivity DNA Kit (#5067-4626, Agilent, Santa Clara, CA). Barcoded libraries were pooled and underwent 75 bp single-end sequencing on an Illumina NextSeq 500.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
processed data file: COUNTS_NeuN.txt
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| Data processing |
Single-end reads were aligned with STAR 2.5.2b using options "“--outFilterMultimapNmax 10 --alignSJoverhangMin 10 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 3 --twopassMode Basic" to a reference sequence (GRCh37 from Ensemble)
RNA-seq Counts values were quantified using HTSeq python suite based on Homo_sapiens.GRCh37.87.gtf from Ensemble based on protein coding genes and exons.
Genome_build: Human hg19 (GRCh37)
Supplementary_files_format_and_content: Input tables and demographics are included as tab separated files.
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| Submission date |
Dec 04, 2017 |
| Last update date |
Nov 15, 2019 |
| Contact name |
Genevieve Konopka |
| E-mail(s) |
gena@alum.mit.edu
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| Organization name |
UT Southwestern Medical Center
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| Department |
Neuroscience
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| Street address |
5323 Harry Hines Blvd.
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| City |
Dallas |
| State/province |
TX |
| ZIP/Postal code |
75390-9111 |
| Country |
USA |
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| Platform ID |
GPL18573 |
| Series (2) |
| GSE107638 |
Contribution of differential cell-specific methylation to schizophrenia [RNA-seq] |
| GSE108066 |
Contribution of differential cell-specific methylation to schizophrenia |
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| Relations |
| SRA |
SRX3440177 |
| BioSample |
SAMN13293159 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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