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Status |
Public on Dec 01, 2022 |
Title |
Cerebellum 175Q 10wk replicate 3 |
Sample type |
SRA |
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Source name |
cerebellum
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Organism |
Mus musculus |
Characteristics |
tissue: cerebellum gender: male age: post natal week 10 genotype: 175Q strain: C57BL/6J
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Extracted molecule |
total RNA |
Extraction protocol |
Collected tissue was stored in RNAlater solution (Thermo Fisher Scientific). Total RNA was isolated using TRIzol reagent (Thermo Fisher Scientific) following the manufacturer’s protocol. Quality control involved quantification using fluorimetry via RiboGreen assay kit (Thermo Fisher Scientific) and RNA integrity was assessed via capillary electrophoresis using an Agilent BioAnalyzer 2100 to generate an RNA integrity number (RIN). Library creation was completed using oligo-dT purification of polyadenylated RNA, which was reverse transcribed to create cDNA. cDNA was fragmented, blunt ended, and ligated to barcode adaptors. Libraries were size selected to 320 bp ± 5% to produce average inserts of approximately 200 bp, and size distribution was validated using capillary electrophoresis and quantified using fluorimetry (PicoGreen, Thermo Fisher Scientific) and qPCR. Libraries were then normalized, pooled, and sequenced on an S4 flow cell by an Illumina NovaSeq 6000 using a 150-nucleotide, paired-end read strategy. Libraries were prepared and sequenced in two batches based on mouse age; 10-week mice were sequenced to a minimum of 70 million reads/sample, while 26-week mice were sequenced to a minimum pf 40 million reads/sample.
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina NovaSeq 6000 |
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Description |
74894_cbm
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Data processing |
Illumina bcl2fastq2 v2.20.0 software used for basecalling. FASTQ files were trimmed, aligned to the mouse reference genome (GRCm38), sorted, and counted using the Bulk RNAseq Analysis Pipeline from the Minnesota Supercomputing Institute’s Collection of Hierarchical UMII/RIS Pipelines (v0.2.0 was used for 26-week-old data. v0.2.2 was used for the 10-week-old data). Component steps for read trimming, aligning and counting are as follows: Reads were trimmed with Trimmomatic v.0.33 using illumina adapters, seedMismatches=4,palindromeClipThreshold=15, simpleClipThreshold=7, minAdapterLength=2 and keepBothReads=true Trimmed reas were aligned to GRCm38 using hisat v.2.1.0 using the parameters --rna-strandedness RF and --no-mixed (suppress unpaired alignments for paired reads). Aligned reads were assinged to genes using the featureCounts function of Subread v.1.6.2 with the parameters -B -p -Q10 -s2 and the Ensembl GRCm38 v.97 annotation gtf. Assembly: GRCm38 Supplementary files format and content: subread_counts_10wk.txt is a matrix of gene counts for each sample at 10 weeks of age Supplementary files format and content: subread_counts_26wk.txt is a matrix of gene counts for each sample at 26 weeks of age
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Submission date |
Nov 18, 2022 |
Last update date |
Dec 01, 2022 |
Contact name |
Christine M. Henzler |
E-mail(s) |
chenzler@umn.edu
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Phone |
6126257889
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Organization name |
University of Minnesota, Twin Cities (Minneapolis, MN, US)
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Department |
Minnesota Supercomputing Institute
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Street address |
117 Pleasant St SE
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City |
Minneapolis |
State/province |
MN |
ZIP/Postal code |
55455 |
Country |
USA |
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Platform ID |
GPL24247 |
Series (1) |
GSE218303 |
Decreasing Mutant ATXN1 Nuclear Localization Improves a Spectrum of SCA1-Like Phenotypes and Brain Region Transcriptomic Profiles |
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Relations |
BioSample |
SAMN31787167 |
SRA |
SRX18308306 |
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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