Rnase A

Ribonuclease A(RNaseA) is a single-stranded polypeptide containing 4 disulfide bonds with a molecular weight of about 13.7 kDa. RNase Ais an endoribonuclease that specifically degrades single-stranded RNA at C and U residues. Specifically,the cleavage recognizes the phosphodiester bond formed by the 5′-ribose of a nucleotide and the phosphate group on the 3′-ribose of the adjacent pyrimidine nucleotide, so that the 2,3′-Cyclic phosphates are hydrolyzed to the corresponding 3′nucleoside phosphates (eg,pG-pG-pC-pA-pG is cleaved by RNase A to generate pG-pG-pCp and A-PG). RNase A is the most active in cleaving single-stranded RNA. Recommended working concentration is 1- 100 μ G/mL, compatible with various reaction systems. Low salt concentration (0-100 mM NaCl) can be used to cut single-stranded RNA, double-stranded RNA, and RNA chains formed by RNA-DNA hybridization.However, at high salt concentration (≥0.3 M),RNase A only specifically cleaves single-stranded RNA.

This is one of the common methods for preparing RNase A storage solution. It can also be

prepared by other methods according to the traditional methods in the laboratory or reference literature (such as

directly dissolving in 10 mM Tris-HCl,pH 7.5 or Tris-NaCl solution)

1. Use 10 mM sodium acetate (pH 5.2)to prepare 10 mg/mL of RNase A storage solution.

2.  Heating at 100 ℃ for 15 min.

3.  Cool to room temperature,add  1/10  volume  of 1 M Tris-HCl  (pH 7.4),adjust its pH to 7.4(for example,add 500 mL of 10 mg/mL RNase storage solution 1 M Tris-HCl,pH7.4).

4.  Sub-packaged at-20℃ for frozen storage,which can be stable for up to 2 years.

[Notes]:

When boiling RNaseA solution under neutral conditions,RNase precipitation will form; Boil it at a lower pH, and if

there is precipitation, it can be observed, which may be caused by the presence of protein  impurities.  If  sediment  is  found  after  boiling,impurities  can  be  removed  by  high-speed

centrifugation (13000    rpm),and    then    sub- packed for freezing storage

Notes

Please wear the necessary PPE, such lab coat and gloves, to ensure your health and safety.