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Cat. No.: MAZF-1k (for 1KU)

Description

MazF is a toxin protein in the E.colitoxin-antitoxin system. It is a specific endonuclease that cleaves single-stranded RNA at ACA sequences at the 5' end. It does not cleave double-stranded RNA, double-stranded DNA, or single-stranded DNA.

Components

• MazF (20 U/μl): 50 μl
• 5× MazF Buffer:  1 ml

Activity Definition

One activity unit (U) is defined as the amount of enzyme required to completely degrade 1 pmol of standard substrate (ROX-5'-GATAUACATATCT-eclipse; the underlined part is the RNA sequence) at 37°C and pH 7.5 within 10 minutes.

Application

• High-resolution m6A methylation analysis;
• mRNA drug sequence analysis.

Quality Control

• Protein Purity: The protein purity is not less than 95% as detected by SDS-PAGE gel electrophoresis.
• RNase Activity: When 20 U MazF is incubated with 40 ng ssRNA at 37°C for 1 hour and detected by agarose gel electrophoresis, there is no change in the ssRNA fragments.
• DNase Activity: When 20 U MazF is incubated with 15 ng double-stranded DNA at 37°C for 16 hours and detected by agarose gel electrophoresis, there is no change in the DNA fragments.

Storage

-20°C.

Precautions

• MazF specifically cleaves ssRNA containing ACA sequences but may also cleave AC sites due to the influence of sequences on both sides of the AC site.
• MazF cannot cleave dsRNA. Therefore, due to the influence of RNA secondary structure, MazF cannot cleave all ACA site sequences.
• Adding salts, especially divalent cations such as Mg2+, to the reaction solution will inhibit MazF activity. DTT does not affect the enzyme activity in the reaction solution.
• Fragments cleaved by MazF cannot be directly ligated using RNA Ligase because the two ends of the fragments are 2', 3'-cyclic phosphate and 5'-OH.