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| Status |
Public on Dec 01, 2021 |
| Title |
BC-1_Mutant_peptide_2 |
| Sample type |
SRA |
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| Source name |
human primary effusion lymphoma
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| Organism |
Homo sapiens |
| Characteristics |
cell line: BC-1
treatment: Mutant peptide (24uM) for 90 minutes
labeling treatment: 4-Thiouridine (s4U; 300 uM) for 1 hour
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| Treatment protocol |
BCBL-1 and BC-1 cell cultures (two cultures per treatment) were incubated with wild type peptide (24 uM), mutant peptide (24 uM), or vehicle control for 30 minutes. Subsequently, 4-Thiouridine (s4U; 300 uM) was added to the culture media and the cells treated for 1 hour in order to label newly synthesized RNA.
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| Growth protocol |
BCBL-1 and BC-1, and BC-3 cell lines were cultured in RPMI 1640 medium supplemented with 15% FBS. The cultures were maintained at 37°C in a humidified atmosphere containing 5% CO2 (v/v). Mycoplasma contamination was routinely tested by using a PCR-based assay.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using the TRIzol Reagent (Invitrogen).
Total RNA was isolated from cells following incubation with 4-Thiouridine (s4U; 300 uM). Subsequently, the 4-thiol groups in the s4Uracil-labeled transcripts were alkylated with iodoacetamide (IAA). QuantSeq 3’ mRNA-Seq (FWD) (Lexogen, Inc.) Illumina-compatible, indexed sequencing libraries were then prepared from the alkylated RNA samples (100 ng) according to the manufacturer’s protocol for oligo(dT)-primed first strand cDNA synthesis, random-primed second strand synthesis, and library amplification.
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 4000 |
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| Description |
Illimuna next-generation sequencing (100bp, single read)
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| Data processing |
Library strategy: SLAM-seq
SLAM-Seq datasets were analyzed using the T>C conversion-aware SLAM-DUNK (Digital Unmasking of Nucleotide conversion-containing k-mers) pipeline utilizing the default parameters (DOI: 10.1038/nmeth.4435, DOI: 10.1186/s12859-019-2849-7). Briefly, nucleotide conversion-aware read mapping of adapter- and poly(A)-trimmed sequences to the human GRCh38/hg38 reference genome assembly was performed with NextGenMap (DOI:10.1093/bioinformatics/btt468). Alignments were filtered for those with a minimum identity of 95% and minimum of 50% of the read bases mapped. For multi-mappers, ambiguous reads and non-3' UTR alignments were discarded, while one read was randomly selected from multimappers aligned to the same 3’ UTR. SNP calling (coverage cut-off of 10X and variant fraction cut-off of 0.8) with VarScan2 (DOI: 10.1101/gr.129684.111) was performed in order to mask actual T>C SNPs prior. Non-SNP T>C conversion events were then counted and the fraction of labeled transcripts determined. All output is contained in tcount files.
Genome_build: GRCh38/hg38
Supplementary_files_format_and_content: The processed data is contained in the tcount file, which are the central output of slamdunk. These contain all results, conversion rates, and other statistics for each UTR. Tcount files are tab-separated text files containing one line entry per annotated 3’ UTR. Each entry contains the following columns: "chromosome, start end, name, length, strand, conversionRate, readsCPM, Tcontent, coverageOnTs, conversionsOnTs, readCount, tcReadCount, and multimapCount.
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| Submission date |
May 09, 2021 |
| Last update date |
Dec 03, 2021 |
| Contact name |
Clifford G. Tepper |
| E-mail(s) |
cgtepper@ucdavis.edu
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| Phone |
916-734-7195
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| Organization name |
UC Davis School of Medicine
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| Department |
Biochemistry and Molecular Medicine
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| Street address |
4645 2nd Avenue, Room 2300A
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| City |
Sacramento |
| State/province |
CA |
| ZIP/Postal code |
95817 |
| Country |
USA |
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| Platform ID |
GPL20301 |
| Series (2) |
| GSE173725 |
Targeting MYC Transcription with Small Peptide Derived from KSHV Transactivator |
| GSE174115 |
Targeting MYC Transcription with Small Peptide Derived from KSHV Transactivator (SLAM-Seq) |
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| Relations |
| BioSample |
SAMN19080777 |
| SRA |
SRX10823550 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5287460_BC1_Mut_2_R1_001_trimmed.fq_slamdunk_mapped_filtered_tcount.tsv.gz |
1.6 Mb |
(ftp)(http) |
TSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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