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. 2003 May 30;100(12):7033–7038. doi: 10.1073/pnas.1232347100

Fig. 2.

Fig. 2.

(A) Comparing 5′ cap- and 3′ poly(A)-dependent purifications of mRNA. Total RNA (315 μg) from normal liver was either mixed with GST-4EK119A beads as described in Materials and Methods or applied to an oligo(dC10T30) column. The GST-4EK119A matrix was washed, and mRNA was recovered by eluting with m7GDP (see Materials and Methods). An oligo(dT) column was used to purify mRNA as suggested by the manufacturer (Qiagen). Ten percent of the mRNA recovered from each purification was analyzed by formaldehyde agarose (1%) gel electrophoresis. An ethidium bromide-stained gel is shown: lane 1, 0.24- to 9.5-kb RNA ladder; lane 2, mRNA batch purified using GST-4EK119A agarose beads; lane 3, mRNA purified using an oligo(dT) column. (B) In vitro translation of mRNA purified using GST-4EK119A or oligo(dT). mRNA (1 μg) isolated by either GST-4EK119A or oligo(dT) was translated in a nuclease-treated rabbit reticulocyte lysate with [35S]methionine (see Materials and Methods). Protein products were analyzed by 10% SDS/PAGE and autoradiography: lane 1, control with no mRNA added; lane 2, mRNA purified with GST-4EK119A; and lane 3, mRNA purified with oligo(dT). Molecular mass standards are shown.