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| Status |
Public on Aug 08, 2021 |
| Title |
104-13 |
| Sample type |
SRA |
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| Source name |
_92.1
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| Organism |
Homo sapiens |
| Characteristics |
cell type: Uveal melanoma cells
cell line: _92.1
treatment: PRT + UMK57
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| Treatment protocol |
Cells were grown under regular conditions until they reached approximately 80% confluence. Culture media (RPMI) containing the following compounds dissolved in DMSO (Fisher BioReagents), or an equal volume of DMSO, were added to the cells; 50 μM PRT (PRT4165, Sigma-Aldrich), 1 μM UMK57 (Aobious) and 0.5 μM reversine (Cayman Chemical). For experiments with treatment extending beyond 2 hours, culture media were supplemented with 10% FBS and replenished after the first 24 hours, with the applicable compound.
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| Growth protocol |
Melanoma cells were grown in RPMI-1640 media (Gibco) supplemented with 10% fetal bovine serum (FBS, Sigma). Cell culture media were supplemented with 1% penicillin/streptomycin and cells were maintained at 37 °C and 5% CO2.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was the extracted using the QIAShredder and the RNeasy extraction kit (Qiagen) and sequenced on NovaSeq (Illumina) to generate 150 bp pair-end reads. CUT&RUN were prepared using the CUTANA kit (EpiCypher, 14-1048)
RNA libraries were prepared for sequencing using standard Illumina protocols, CUT&RUN Library preparation was performed using the NEBNext Ultra II Library Prep Kit for Illumina (New England BioLabs, E7103S)
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
Exp2_Gene_normalized_counts.txt
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| Data processing |
Raw reads (FASTQ) were aligned to the human reference genome (Ch38) using STAR on Partek Flow software, version 8.0. Aligned reads were mapped to transcripts (Ensembl Transcripts release 102) using Partek’s modified expectation-maximization (EM) algorithm for transcript quantification, with strict paired-end compatibility. Gene counts were normalized per sample to generate fragments Per Kilobase of transcript per Million mapped reads (FPKM) values.
Raw reads (FASTQ) were aligned to the human reference genome (Ch38), using STAR on Partek Flow software, version 10.0 (Partek). Genomic peaks were called using MACS2 with the following parameters (BAM, cutoff q-value 0.05, no down-sampling).
Genome_build: hg38
Supplementary_files_format_and_content: Matrix table with normalized counts for every gene and every sample
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| Submission date |
Aug 06, 2021 |
| Last update date |
Aug 08, 2021 |
| Contact name |
Mathieu F Bakhoum |
| Organization name |
Yale University
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| Department |
Ophthalmology
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| Lab |
Bakhoum Laboratory
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| Street address |
300 George St.
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| City |
New Haven |
| State/province |
CT |
| ZIP/Postal code |
06510 |
| Country |
USA |
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| Platform ID |
GPL24676 |
| Series (1) |
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| Relations |
| BioSample |
SAMN20603299 |
| SRA |
SRX11664680 |
| 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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