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| Status |
Public on Feb 19, 2024 |
| Title |
Microglia, replicate 1, batch 2 |
| Sample type |
SRA |
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| Source name |
Cortex
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| Organism |
Mus musculus |
| Characteristics |
tissue: Cortex
cell type: Cortical microglia
genotype: Rpl22 Fl/+, Tmem119-cre
Sex: Female
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| Growth protocol |
Female B6N.129-Rpl22tm1.1Psam/J mice were crossed to the appropriate Cre driver lines to generate Rpl22 fl/+, CreTg/+ mice. Tissue was harvested from 6-week-old mice
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
Freshly isolated tissue was homogenized on ice using a dounce homogenizer in lysis buffer (50mM Tris-Cl pH 7.5, 100mM KCl, 12mM MgCl2, 1mM DTT, 100μg/ml cycloheximide, 1% (v/v) NP-40, 1mg/ml heparin, 25 units/ml Turbo DNaseI (ThermoFisher Scientific), 20 units/ml Superase-In RNAse inhibitor (ThermoFisher Scientific), cOmplete mini protease inhibitor cocktail (Roche)). Ribosomes were then isolated as previously described, with some minor modifications (Sanz et al., 2009). Briefly, for each sample, 200 ml of Pierce anti-HA magnetic beads (ThermoFisher Scientific) was washed in lysis buffer and then added to the lysate containing tagged ribosomes. The samples were incubated for 4 hours at 4ºC with rotation. Ribosome-bound beads were then washed 3 times for 5 minutes each in 1 ml of high salt wash buffer (50mM Tris pH 7.4, 300mM KCl, 12mM MgCl2, 0.5mM DTT, 100μg/ml cycloheximide, 1 % (v/v) NP-40, 20 units/ml Superase-In RNAse inhibitor, cOmplete mini protease inhibitor cocktail). After the final wash, the ribosome-bound RNA was eluted using buffer RLT (supplemented with b-mercaptoethanol) and isolated using the Qiagen RNeasy mini kit following manufacturer’s instructions. RNA was eluted in 30 μL nuclease-free H2O and the quality and concentration were assessed using high sensitivity RNA ScreenTape on a TapeStation (Agilent) and the Qubit RNA high sensitivity kit on a Qubit 2.0 fluorometer (Life Technologies).
Three mRNA libraries were generated for each genotype using the Kapa Stranded mRNA-Seq Kit (Roche) according to the manufacturer’s protocol.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Reads were trimmed to remove adapter sequences (AGATCGGAAGAG) using cutadapt version 3.5 (Martin, 2011) and then quantified using kallisto version 0.46.2 (Bray et al., 2016) pseudo-aligning to a Gencode M24 transcript reference.
Assembly: Gencode M24 transcript
Supplementary files format and content: Tab delimited text files include counts and tpm for the sample
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| Submission date |
Aug 01, 2023 |
| Last update date |
Feb 19, 2024 |
| Contact name |
Mridu Kapur |
| E-mail(s) |
mkapur@health.ucsd.edu
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| Organization name |
University of California San Diego
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| Department |
Cellular and Molecular Medicine
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| Lab |
Susan Ackerman
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| Street address |
9500 Gilman Drive
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| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92093 |
| Country |
USA |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE239759 |
Cell-type-specific expression of tRNAs in the brain regulates cellular homeostasis |
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| Relations |
| BioSample |
SAMN36781006 |
| SRA |
SRX21206896 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7671090_Tmem_1_abundance.tsv.gz |
4.5 Mb |
(ftp)(http) |
TSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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