{"doi":"10.1261/rna.079776.123","title":"SCARPET: site-specific quantification of methylated and nonmethylated adenosines reveals m<sup>6</sup>A stoichiometry","abstract":"m 6 A has different stoichiometry at different positions in different mRNAs. However, the exact stoichiometry of m 6 A is difficult to measure. Here, we describe SCARPET (site-specific cleavage and radioactive-labeling followed by purification, exonuclease digestion, and thin-layer chromatography), a simple and streamlined biochemical assay for quantifying m 6 A at any specific site in any mRNA. SCARPET involves a site-specific cleavage of mRNA immediately 5′ of an adenosine site in an mRNA. This site is radiolabeled with 32 P, and after a series of steps to purify the RNA and to remove nonspecific signals, the nucleotide is resolved by TLC to visualize A and m 6 A at this site. Quantification of these spots reveals the m 6 A stoichiometry at the site of interest. SCARPET can be applied to poly(A)-enriched RNA, or preferably purified mRNA, which produces more accurate m 6 A stoichiometry measurements. We show that sample processing steps of SCARPET can be performed in a single day, and results in a specific and accurate measurement of m 6 A stoichiometry at specific sites in mRNA. Using SCARPET, we measure exact m 6 A stoichiometries in specific mRNAs and show that Zika genomic RNA lacks m 6 A at previously mapped sites. SCARPET will be useful for testing specific sites for their m 6 A stoichiometry and to assess how m 6 A stoichiometry changes in different conditions and cellular contexts.","journal":"RNA","year":2023,"id":370477,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":7,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9575,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2023-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":109224,"name":"Yaron Bram","orcid":"0000-0001-9711-575X","position":1,"is_corresponding":false},{"id":103718,"name":"Robert E. Schwartz","orcid":"0000-0002-5417-5995","position":2,"is_corresponding":false},{"id":107966,"name":"Samie R. Jaffrey","orcid":"0000-0003-3615-6958","position":3,"is_corresponding":false},{"id":415476,"name":"Aashiq H. Mirza","orcid":"0000-0002-9655-5267","position":0,"is_corresponding":true}],"reference_count":46,"raw_metadata":null,"created_at":"2026-07-19T01:15:37.094100Z","pmid":"38190635","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}