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. Author manuscript; available in PMC: 2015 Jun 23.
Published in final edited form as: Methods Enzymol. 2015 Feb 27;558:3–37. doi: 10.1016/bs.mie.2015.01.008

Figure 1.

Figure 1

Enzymatic synthesis and purification of long noncoding RNAs. (A) The native purification pipeline of lncRNAs comprises the following steps (from left to right and top to bottom): the RNA is synthesized with the T7 RNA polymerase system; when the transcription reaction is finished, the DNA template is digested with DNase enzyme; enzymes in the reaction are then proteolyzed with the action of the proteinase K enzyme; RNA is captured from reaction components and buffer exchanged by ultra-filtration with Amicon centrifugal devices (Millipore); finally, the RNA is subjected to size-exclusion chromatography. (B) The resulting chromatogram represents the absorbance of the filtered RNA as a function of the elution volume. Material in the void volume of the column (10 ml elution) and shorter RNA molecules (22 ml elution) are excluded. The fraction(s) corresponding to the main peak are then selected for downstream analysis.