Figure 4.

Poly(A) mRNA enrichment and m³C levels monitored during purification. A, schematics of poly(A) mRNA enrichment and separation of 28S and 18S rRNA. B, typical electropherogram for the following: 1. total RNA; 2. RNA >5.8S isolated by SEC; 3. poly(A) RNA enriched by oligo(dT) selection; 4. poly(A)-enriched mRNA after rRNA depletion; 5. 28S; and 6. 18S rRNA purified by SEC. C, LC-MS/MS quantitation of m³C level in tRNA, RNA larger than 5.8S rRNA, oligo(dT)-enriched mRNA, ribo-zero-treated mRNA, 18S rRNA, 28S rRNA. D, LC-MS/MS quantitation of m³C, m⁵C, m¹A, and m⁶A in human HCT116 mRNA purified by SEC, poly(A)-enrichment, and rRNA depletion. (The level of each modified ribonucleoside is expressed relative to its unmodified form. Data represent mean ± S.D. for n > = 3.)