title: Probing N⁶-methyladenosine (m⁶A) RNA Modification in Total RNA with SCARLET.
abstract: Posttranscriptional N (6)-methyladenosine (m(6)A) RNA modification is indispensable for cell development and viability; however, functional investigation of m(6)A biological function has been hindered by the lack of methods for its precise identification and quantitation. Here, we describe a method that accurately identifies m(6)A position and modification fraction in human messenger RNA (mRNA) and long noncoding RNA (lncRNA) at single-nucleotide resolution, termed as "site-specific cleavage and radioactive-labeling followed by ligation-assisted extraction and thin-layer chromatography (SCARLET)" (Fig. 1). This method combines two previously established techniques, site-specific cleavage and splint ligation, to probe the m(6)A RNA modification status at any mRNA/lncRNA site in the total RNA pool.
authors.affiliation: Department of Chemistry, University of Chicago, Chicago, IL, 60637, USA.
authors.forename: Nian
authors.initials: N
authors.lastname: Liu
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, IL, 60637, USA. taopan@uchicago.edu.|Institute of Biophysical Dynamics, University of Chicago, Chicago, IL, 60637, USA. taopan@uchicago.edu.
authors.forename: Tao
authors.initials: T
authors.lastname: Pan
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives* / Q000235:genetics; D002855:Chromatography, Thin Layer / Q000379:methods*; D006367:HeLa Cells; D006801:Humans; D008745:Methylation; D012323:RNA Processing, Post-Transcriptional / Q000235:genetics*; D062085:RNA, Long Noncoding / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D011868:Radioisotopes / Q000737:chemistry
chemical_list: D062085:RNA, Long Noncoding; D012333:RNA, Messenger; D011868:Radioisotopes; C010223:N-methyladenosine; D000241:Adenosine
keywords: Modification fraction; N 6-methyladenosine; RNA modification; SCARLET; Single-nucleotide resolution; mRNA /lncRNA