Skip to main content
. 2019 Feb 27;15:26–35. doi: 10.1016/j.omtn.2019.02.018

Figure 1.

Figure 1

Evaluating the Purity of IVT mRNA following Cellulose Chromatography

(A) Uridine (U)- or 1-methylpseudouridine (m1Ψ)-containing, 2.2-kb-long IVT mRNA present in 16% ethanol-containing buffer was purified using cellulose-filled microcentrifuge spin columns. A dot blot loaded with aliquots of mRNA was analyzed for the presence of dsRNA contaminants with J2 mAb and reprobed with complementary oligodeoxynucleotide (ODN). The corresponding mRNA fractions were analyzed by electrophoresis on a 1.4% agarose gel. The GelRed-stained RNAs were visualized by UV illumination. (B) In vitro-transcribed mRNA containing m1Ψ was purified repeatedly in three consecutive cycles using cellulose-filled microcentrifuge spin columns. Aliquots of mRNA were analyzed in dot blot with J2 dsRNA-specific mAb and S9.6 mAb specific for RNA:DNA hybrids. The mRNA fractions separated in 1.4% agarose gel were stained with GelRed and visualized by UV illumination. (C) Linearized plasmid and U-containing dsRNA were mixed in 16% ethanol-containing buffer and then subjected to cellulose chromatography. DNA recovered from the unbound fraction and dsRNA eluted from the cellulose-bound fraction were separated on 1% agarose gel, then stained with GelRed and visualized by UV illumination. M, DNA molecular weight marker.