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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jul 15, 2019 |
| Title |
PR3 BTSC |
| Sample type |
SRA |
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| Source name |
BTSC_1
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| Organism |
Mus musculus |
| Characteristics |
tissue: brain
cell type: mouse brain tumor stem cells
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| Extracted molecule |
total RNA |
| Extraction protocol |
The cells of a 15 cm culture plate were lysed with lysis buffer (20 mM Tris-HCl pH 7.4, 10 mM MgCl2, 200 mM KCl, 1 % Triton, 2 mM DTT, 100 µg/ml Cyclohexamide, 25 U/ml DNase and EDTA-free protease inhibitors, Roche). The mRNA not protected by ribosomes was digested with RNAseI and the ribosome footprints were collected by centrifugation with a continuous sucrose gradient (10-50 %) with DTT, cyclohexamideand 20 U/ml SUPERaseIN. The fraction with the ribosome footprint RNA was used for acid phenol extraction.
Precipitated RNA was resuspended in 10 mM Tris pH7, mixed with 2x sample buffer (TBE-urea, NOVEX, Invitrogen) and loaded on a 15 % TBE-urea gel. The region between 26-34 nt was excised and used for library generation exactly as described previously (Ingolia et al, Nat Protoc 2012). The library was sequenced on the Illumina HiSeq system according to the manufacturer’s protocol.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
ribosome protected mRNA
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| Data processing |
Bowtie2 was used to align the reads against mouse rRNA sequences.
Only the non-aligned reads were used for further analysis with TopHat v1.4.0 using the reference genome mm10.
To analyse the differentially expressed genes between we used Cuffdiff v2.2.1 with classic fpkm as library normalization method and a FDR of 0.1.
Genome_build: mm10
Supplementary_files_format_and_content: deferentially expressed genes between BTSCs and NSCs under the cut-off of FDR < 0.1
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| Submission date |
Feb 20, 2018 |
| Last update date |
Jul 15, 2019 |
| Contact name |
Haikun Liu |
| E-mail(s) |
l.haikun@dkfz.de
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| Organization name |
DKFZ
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| Street address |
INF 581
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| City |
Heidelberg |
| ZIP/Postal code |
69120 |
| Country |
Germany |
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| Platform ID |
GPL13112 |
| Series (2) |
| GSE110866 |
Identification and validation of GPD1 as an essential cancer stem cell specific target in glioblastoma (Ribosome profiling) |
| GSE110869 |
Identification and validation of GPD1 as an essential cancer stem cell specific target in glioblastoma |
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| Relations |
| BioSample |
SAMN08567970 |
| SRA |
SRX3723693 |
| 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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