Detection and Quantification of N6-Methyladenosine in Messenger RNA by TLC
The base-modified nucleotide, N6-methyladenosine, is a relatively abundant modification found in the mRNA of most higher eukaryotes. Methylation levels can change dependent upon environmental conditions, cell differentiation state, or following knockdown of members of the methylase complex, and it is often useful to directly measure and compare N6-methyladenosine levels between samples. Two dimensional chromatography of radiolabeled nucleotides, following specific nuclease treatments, provides a robust, sensitive, and reproducible assay for this modification.
Key wordsN6-Methyladenosine m6A Thin Layer Chromatography TLC mRNA Methylation Epitranscriptome
- 11.Sambrook J, Russel DW (2001) Molecular cloning : a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp. 7.13–7.17Google Scholar
- 12.Conrad RC, AMBION, INC. High efficiency mRNA isolation. United States Patent Application Publication, Pub. No.: US 2004/0230048 A1 Conrad Pub. Date: 18 Nov 2004.Google Scholar