RNA 5’ Polyphosphatase
- Catalyzes the removal of γ and β phosphate groups from
5'-triphosphorylated and 5'- diphosphorylated RNA, resulting in
5’-monophosphorylated RNA. - Two protocols provided for easy- and hard-to-treat samples.
- Use in conjunction with XRN1 to remove uncapped RNA from
mRNA preparations. - Use for 5’ end RNA substrate preparation for
T4 RNA Ligase-based reactions.
Product is for research use only (RUO)
Product Description:
RNA 5' Polyphosphatase is a patented enzyme that catalyzes the removal of γ and β phosphate groups from 5'-triphosphorylated and 5'-diphosphorylated RNA resulting in 5'-monophosphorylated RNA. Capped RNA is not a substrate for the enzyme.
RNA 5' Polyphosphatase can be used for the following applications:
- Removal of uncapped RNA from an mRNA preparation, in conjunction with XRN1 treatment.
- 5' end RNA substrate preparation for RNA Ligase-based circularization reactions.
- 5' end RNA substrate preparation for T4 RNA Ligase-based reactions (end tagging).
RNA 5' Polyphosphatase is active on nucleoside triphosphates and is inhibited by inorganic phosphate, a natural reaction product. Removal of unincorporated nucleotides and inorganic phosphates from completed transcription reactions is recommended for full enzymatic activity on the intended RNA substrate.
Product Performance:
Figure 1. Two different 5'-triphosphate primary transcript RNAs were treated with RNA 5' Polyphosphatase using the Standard Protocol (no pre heat denaturation step used). After the 30 minute incubation, additional MgCl2 (for buffer compatibility) and XRN1 (5'-monophosphate-dependent exonuclease) was added to the reaction to digest the RNA 5' Polyphosphatase reaction product. Following treatment with RNA 5’ Polyphosphatase and XRN1, the 1.85-kb RNA (lane 2) and 0.92-kb RNA (lane 4) were digested.
Figure 2. A 1.57-kb 5'-triphosphate primary transcript RNA containing stable secondary structure on the 5' end was treated with RNA 5' Polyphosphatase using the Alternate Protocol (pre heat denaturation step used). After the 30 minute incubation, additional MgCl2 (for buffer compatibility) and XRN1 (5'-monophosphate-dependent exonuclease) was added to the reaction to digest the RNA 5' Polyphosphatase reaction product. Heat denaturation prior to RNA 5’ Polyphosphatase and XRN1 treatment improved reaction efficiency (lane 6).
