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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Dec 11, 2019 |
| Title |
37_L1.N701 |
| Sample type |
SRA |
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| Source name |
MMTV-PyMT mammary tumors
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| Organism |
Mus musculus |
| Characteristics |
cell type: CD31+ cells
treatment: A2V+CD40
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| Treatment protocol |
When tumors reached a size of 300 mm3, mice were injected i.p with the following moAbs: mouse IgG1 (clone MOPC-21, 500 μg) + rat IgG2a (clone 2A3, 100 μg), designated as “IgG”; or anti-VEGFA/ANGPT2 (A2V, 500 μg) + rat IgG2a (clone 2A3, 100 μg), designated “A2V”. Five days later, mice were euthanized and tumors harvested and prepared for fluorescence-activated cell sorting (FACS).
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| Extracted molecule |
total RNA |
| Extraction protocol |
Cell sorting was performed using FACS Aria II apparatus (BD). Each cell population (n=4) was isolated from an independent tumor. Typically, 2x104 to 2x105 cells were obtained for each cell population. Sorted cell populations were immediately homogenized in 700 µl Qiazol reagent (Qiagen). 140 µl of chloroform was added to the homogenate and the mixture centrifuged at 16,000 x g for 5 min at 4°C in Phase Lock Tubes (Qiagen). RNA was subsequently extracted from the aqueous phase using the RNeasy Micro kit (Qiagen) according to manufacturer’s instructions. Genomic DNA was removed using the RNase free DNase set (Qiagen) during the extraction. RNA was quality controlled on Agilent Eukaryote Total RNA pico chips (Agilent Technologies). High quality RNA (RIN >7) was obtained for all samples. SMARTer ultra low RNA kit for Illumina sequencing (Clontech) was used to prepare and amplify cDNA from 520 pg of total RNA according to manufacturer’s instructions.
Then, 1 ng of amplified cDNA was subjected to Nextera XT library preparation (Illumina) according to manufacturer’s instructions. Sequencing libraries were quantified using the Kapa Library Quantification kit (Kapa Biosystems) and quality controlled by capillary electrophoresis on a Bioanalyzer using High Sensitivity chips (Agilent Technologies). Libraries were sequenced on a HiSeq2500 sequencer (Illumina) for 2 x 50 cycles using version 4 cluster generation kits and version 4 sequencing reagents (Illumina).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Raw read data was aligned against mouse transcriptome (MouseEnsembl release 61) using Bowtie2 (Langmead and Salzberg, 2012). Splice junctions were defined as available in Ensembl v75. Read mappings were then mapped to genes and transcripts based on the transcript models annotated in Ensembl v75 using in-house tools and, again using in-house software, RPKM values were computed as described previously (Mortazavi et al., 2008).
Genome_build: MouseEnsembl release 61
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each sample
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| Submission date |
Dec 10, 2019 |
| Last update date |
Dec 11, 2019 |
| Contact name |
Chia-Huey Ooi |
| Organization name |
F. Hoffmann-La Roche Ltd.
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| Street address |
Grenzacherstrasse 124
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| City |
Basel |
| ZIP/Postal code |
4070 |
| Country |
Switzerland |
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| Platform ID |
GPL17021 |
| Series (1) |
| GSE94920 |
Molecular analysis of endothelial and immune cells after anti-angiogenic cancer immunotherapy |
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| Relations |
| BioSample |
SAMN13526832 |
| SRA |
SRX7320000 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4213135_37_L1.N701.expression.txt.gz |
1.0 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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