E. coli Ribonuclease H (RNase H) is an endonuclease that specifically degrades the RNA of DNA-RNA hybrids, without affecting DNA or unhybribized RNA. This highly purified preparation is suited for use in RNA Amplification reactions. This enzyme preparation is prepared in a storage buffer containing 1.0 M Trehalose. Please inquire for custom concentrations and bulk quantities. Applications: Enabling the synthesis of second-strand cDNA by removal of the RNA Used in conjunction with AMV RT and T7 RNA Polymerase in amplification of RNA Unit Definition: One unit of RNaseH is that amount of enzyme which catalysis the production of one nmol of acid-soluble nucleotide in 20 minutes at 37oC using the following reaction conditions: 40 mM TrisHCl, pH 7.5 1.0 米M [3H]-poly (rA):24 米M poly(dT) 4.0 mM MgCl2 1.0 mM DTT 30 米g/mL BSA 4.0% glycerol Storage Buffer Conditions of RNaseH in Glycerol: 20:0 mM TrisHCl, pH 7.5 300 mM KCl 20.0 mM Magnesium Acetate 7.0 mM EDTA 1.0 mM Dithiothreitol 50% Glycerol 0.2% Triton X100 Storage Buffer Conditions of RNaseH in Trehalose. Same as above, except for 1.0 M Trehalose instead of Glycerol. Quality Control: DNase Activity: One-half 米g of Hae fragments of Phi X-174 DNA is incubated at 37oC with 2.5 units of RNase H for 3 hours, and then electrophoresed in a native agarose gel simultaneously with control positive DNase 1 reactions. No more than the equivalent of 2.5 X 10 E-4 unit of DNase 1 is detected. Ribonuclease Activity: One microgram of an RNA Ladder is incubated for 2 hours at 37oC with 4.0 units of RNase H, and then electrophoresed in a native agarose gel simultaneously with control positive RNase 1A reactions. No more than the equivalent of 8 X 10-8 unit of RNase 1A is detected. Specific Activity: The specific activity of the E. coli RNase H is no less than 300,000 units per mg. References: Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, (2nd Ed.), 8.64 每 8.65