Sp6 scribe standard rna ivt kit
The SP6-Scribe Standard RNA IVT Kit is a laboratory product designed for in vitro transcription (IVT) of RNA. It contains the necessary components to synthesize RNA from a DNA template using the SP6 RNA polymerase.
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8 protocols using sp6 scribe standard rna ivt kit
Synthesis of Radiolabeled dsRNA Substrates
Capped and Polyadenylated mRNA Synthesis
Radioactive GFP Fragment Synthesis
Synthesis and Purification of Radiolabeled RNA
In Vitro Synthesis of Reporter mRNAs
Reporter mRNAs for in vitro translation experiments were transcribed with a SP6-Scribe Standard RNA IVT Kit (CellScript). Polyadenylated YB-3_Fluc_YB-3 mRNA was transcribed from pSP36T-5ʹUTR_YB-3-FLuc-3ʹUTR_YB-3-A50 linearized with HpaI. Polyadenylated beta-globin_Fluc mRNA was transcribed from pSP36TLuc-A50 linearized with SmaI. mRNAs were capped using the ScriptCap m7G Capping System and the ScriptCap 2ʹ-O-Methyltransferase Enzyme (CellScript) according to the manufacturer’s manual.
Capped and m6A-Containing mRNA Synthesis
To obtain m6A-containing mRNA for experiments on the formation of 48S complexes, ATP was completely replaced by m6ATP at the same concentration (4 mM) in the transcriptional mixture. For translation in a cell-free system, mRNA containing 50% m6A was used. To obtain such mRNA, a mixture of 2 mM ATP and 2 mM m6ATP was used for in vitro transcription. The quality of the obtained mRNAs was checked by electrophoresis in 6% polyacrylamide gel (PAGE) containing 7 M urea.
In vitro dsRNA Processing Assay
Synthesis and Injection of Capped mRNAs
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