Nuclease cleaves RNA at a specific site
Nuclease is a useful tool for obtaining site-specific cleavage products of long fragments of RNA for the study of RNA modification, for modification site analysis, and for obtaining RNA transcription products with indistinguishable length ends.
Operation method
Nuclease cleaves RNA at a specific site
Materials and Instruments
Synthesized nuclease molecule Cutting substrate RNA Move I. Materials and equipment Caveat 1) Cutting substrate RNA and nuclease can be obtained by in vitro transcription.2) The cutting substrate RNA should be matched with the nuclease. If the substrate and the nuclease have longer base pairing, the specificity of cleavage can be improved, but the number of cleavage transitions will be reduced, so generally 5-7 nucleotides are chosen as the complementary matching region.3) In order to obtain the best cleavage effect, the amount of substrate RNA, nuclease, concentration of magnesium ion, reaction temperature and reaction time should be determined by experiment. For more product details, please visit Aladdin Scientific website.
Tris-HCl MgCl2
1) 500 mmol/L Tris-HCl (pH 7.4)
2)100 mmol/LMgCl2
3) Synthesized nuclease molecule
4) Cutting of substrate RNA
II. Methods of Operation
1) Preparation of cutting mixture: no more than l0 pmol of substrate RNA, l pmol of synthesized nuclease molecule, add water to 8 ul, add l ul 500 mmol/L Tris-HCl (PH7.4), mix well.
2) Heat at 75℃ for 1 min, then place on ice.
3) Add 1ul of 100 mmol/L MgCl2 and incubate at 42℃ for 1h.
4) Separate by denaturing polyacrylamide gel electrophoresis and recover.