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| Status |
Public on Mar 03, 2023 |
| Title |
P252.2-T-RNAseq |
| Sample type |
SRA |
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| Source name |
Tissue; Patient P252.2 tumor; Liposarcoma
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| Organism |
Homo sapiens |
| Characteristics |
tissue: Patient P252.2 tumor
disease state: Liposarcoma
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| Treatment protocol |
Patient liposarcoma samples were ground in liquid nitrogen, transferred to dounce homogenizer and lyzed in cold lysis buffer (20mM Tris-HCl, pH 8.0, 15 mM NaCl, 60 mM KCl, 320 mM sucrose, 1 mM DTT, 0.1% triton X-100, 0.1 mM PMSF, 1x proteinase inhibitor cocktail) with douncing around 20 times using pestle B on the ice. Then the supernatant containing the nuclei was transferred to a 15 ml tube avoiding pipetting tissue debris. Nuclei were pelleted by spinning 6 min at 600 g in a swing-bucket centrifuge and were resuspended in lysis buffer. The same volume of 1.4 M sucrose buffer (20mM Tris-HCl, pH 8.0, 15 mM NaCl, 60 mM KCl, 1.4 M sucrose, 1 mM DTT, 0.1 mM PMSF, 1x proteinase inhibitor cocktail) was carefully added to the bottom of 15 ml tube underneath of nuclei solution. Then, the nuclei were purified by spinning 30 min at 2500 g in swing-bucket centrifuge and resuspended in 200 ul of 320 mM sucrose buffer (20mM Tris-HCl, pH 8.0, 15 mM NaCl, 60 mM KCl, 320 mM sucrose, 1 mM DTT, 0.1 mM PMSF, 1x proteinase inhibitor cocktail).
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
20 mg of ground patient samples or 1.5 million liposarcoma cells were used for RNA-seq. The total RNA was extracted from Trizol according to the manufacturer’s protocol (Invitrogen). PolyA RNA was purified from 1,000 ng of total RNA using oligo (dT) beads (Invitrogen).
Extracted RNA was first fragmented, then followed by reverse transcription, end-repair, adenylation, adaptor ligation, and subsequent PCR amplification. The final product was checked by size distribution and concentration using a BioAnalyzer High Sensitivity DNA Kit (Agilent) and Kapa Library Quantification Kit (Kapa Biosystems).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
Tissue
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| Data processing |
RNA-seq reads were aligned to hg38 genome assembly using STAR; The TPM value of gene expression was caculated using RSEM.
ChIP-seq and ATAC-seq reads were aligned to hg38 genome assembly using Bowtie2;H3K27ac ChIP-Seq peaks were called using MACS2.
Hi-C raw FASTQ files and then mapped the trimmed files against hg38 human reference genome using runHiC pipeline. Hi-C matrix was generated using cooler.
WGS raw FASTQ files were first trimmed for adapter and then mapped against hg38 human reference genome using BWA-MEM algorithm (0.7.17-r1188). Duplicate reads were removed using Picard Tools.
Nanopore raw FASTQ files were mapped against the hg38 reference genome using the minimap2. The structure variations were identified using Sniffles.
Assembly: hg38 (GRCh38)
Supplementary files format and content: tab-delimited text files include TPM values for each Sample; the bigwig files included sequencing reads enrichment signal for each Sample; The .cool file were the multi-resolution matrix of Hi-C for each Sample; the .vcf files included the structure variation information.
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| Submission date |
Apr 19, 2022 |
| Last update date |
Mar 03, 2023 |
| Contact name |
Feng Yue |
| E-mail(s) |
yue@northwestern.edu
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| Organization name |
Northwestern University Feinberg School of Medicine
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| Street address |
303 E. Superior Simpson Querrey
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| City |
CHICACO |
| State/province |
ILLINOIS |
| ZIP/Postal code |
60611 |
| Country |
USA |
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| Platform ID |
GPL24676 |
| Series (1) |
| GSE201056 |
Integrated epigenomic analysis identifies structural alterations associated with oncogenic gene expression in liposarcoma |
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| Supplementary file |
Size |
Download |
File type/resource |
| GSM6049709_P252.2-T.tpm.txt.gz |
259.0 Kb |
(ftp)(http) |
TXT |
| Processed data provided as supplementary file |
| Raw data not provided for this record |
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