github link
Accession IconSRP109095

Thiol-linked alkylation for the metabolic sequencing of RNA [SLAM-seq of wildtype mES cell RNA +/- iodoacetamide treatment]

Organism Icon Mus musculus
Sample Icon 6 Downloadable Samples
Technology Badge IconIllumina HiSeq 2500

Submitter Supplied Information

Description
Gene expression profiling by high-throughput sequencing reveals qualitative and quantitative changes in RNA species at steady-state but obscures the intracellular dynamics of RNA transcription, processing and decay. We developed thiol(SH)-linked alkylation for the metabolic sequencing of RNA (SLAM-seq), an orthogonal chemistry-based epitranscriptomics-sequencing technology that uncovers 4-thiouridine (s4U)-incorporation in RNA species at single-nucleotide resolution. In combination with well-established metabolic RNA labeling protocols and coupled to standard, low-input, high-throughput RNA sequencing methods, SLAM-seq enables rapid access to RNA polymerase II-dependent gene expression dynamics in the context of total RNA. When applied to mouse embryonic stem cells, SLAM-seq provides global and transcript-specific insights into pluripotency-associated gene expression. We validated the method by showing that the RNA-polymerase II-dependent transcriptional output scales with Oct4/Sox2/Nanog-defined enhancer activity; and provides quantitative and mechanistic evidence for transcript-specific RNA turnover mediated by post-transcriptional gene regulatory pathways initiated by microRNAs and N6-methyladenosine. SLAM-seq facilitates the dissection of fundamental mechanisms that control gene expression in an accessible, cost-effective, and scalable manner. Overall design: Total RNA from wildtype mouse embryonic stem (mES cells) was extracted and subjected to alkylation or mock treatment prior to mRNA 3' end library preparation (QuantSeq, Lexogen) and high throughput sequencing.
PubMed ID
Total Samples
6
Submitter’s Institution
No associated institution

Samples

Show of 0 Total Samples
Filter
Add/Remove
Accession Code
Title
Specimen part
Treatment
Subject
Processing Information
Additional Metadata
No rows found
Loading...