|
| Status |
Public on Jun 23, 2024 |
| Title |
IWR4, replicate1, mRNASeq |
| Sample type |
SRA |
| |
|
| Source name |
Brain Tumor
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Brain Tumor
cell type: Leukocytes
diagnosis: Glioblastoma
genotype: IDH Wild Type
primary/recurrent: Recurrent
treatment: Standard of care
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Illumina Compatible low input mRNA libraries were prepared using the Smart-Seq V4 Ultra Low Input RNA kit (Takara Bio, USA) and KAPA HyperPlus Library Preparation kit (Roche). Briefly, full length, double-stranded cDNA was generated from 8ng of total RNA using Takara’s SMART (Switching Mechanism at 5’ end of RNA Template) technology. The ds cDNA was amplified by nine cycles of LD-PCR, then purified using Ampure Beads (Agencort). Following bead elution, the cDNA was evaluated for size distribution and quantity using the Fragment Analyzer High Sensitivity NGS Fragment Analysis Kit (Agilent Technologies) and the Qubit dsDNA HS Assay Kit (ThermoFisher) respectively. The cDNA was enzymatically fragmented, and 20ng of the fragmented cDNA was used to generate Illumina compatible libraries using the KAPA HyperPlus Library Preparation kit. The KAPA libraries were purified and enriched with 2 cycles of PCR to create the final cDNA library. The libraries were quantified using the Qubit™ dsDNA HS Assay (ThermoFisher), then multiplexed 7 libraries per pool. The pooled libraries were quantified by qPCR using the KAPA Library Quantification Kit (KAPA Biosystems), and assessed for size distribution using the TapeStation 4200 (Agilent Technologies). The libraries were then sequenced, one pool per lane, on the Illumina HiSeq4000 sequencer using the 76bp paired end format.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Data processing |
STAR 2-pass alignment (v2.7.0f) was performed with default parameters to generate RNA- seq BAM files.
HTSeq-count (v0.11.0) tool was applied to aligned RNA-seq BAM files to count for each gene how many aligned reads overlap with its exons
RNA-Seq gene expression raw read counts generated from HTSeq-count (v0.11.0) were normalized into Transcripts Per Kilobase Million (TPM).
Assembly: hg19
Supplementary files format and content: tab-delimited text files include read count of each gene for each Sample
|
| |
|
| Submission date |
Jan 10, 2023 |
| Last update date |
Jun 23, 2024 |
| Contact name |
Jeffrey Andrew How |
| E-mail(s) |
jahow@mdanderson.org
|
| Organization name |
The University of Texas MD Anderson Cancer Center
|
| Department |
Gyn Onc & Reproductive Med
|
| Street address |
1515 Holcombe Blvd. Unit 1362
|
| City |
Houston |
| State/province |
TX |
| ZIP/Postal code |
77030-4009 |
| Country |
USA |
| |
|
| Platform ID |
GPL20301 |
| Series (2) |
| GSE222518 |
Immune landscape of Isocitrate Dehydrogenase (IDH) stratified human gliomas [RNA-Seq] |
| GSE222522 |
Immune landscape of Isocitrate Dehydrogenase (IDH) stratified human gliomas |
|
