Ribonuclease H from E.coli
Ribonuclease H is an endonuclease which specifically degrades the RNA moiety of a DNA:RNA hybrid, without affecting the unhybridized RNA. It does not affect double-stranded RNA or DNA. It is provided in a 50% glycerol storage buffer. It is also available in a Trehalose Buffer (Suitable for lyophilization). Concentration 1000 U/mL.
Please inquire for custom concentrations and bulk quantities.
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
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
[3H]-poly (rA):24 µM poly(dT)
4.0 mM MgCl2
1.0 mM DTT
30 µg/mL BSA
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
0.2% Triton X100
Storage Buffer Conditions of RNaseH in Trehalose. Same as above, except for 1.0 M Trehalose instead of Glycerol.
One-half µg 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.
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.
The specific activity of the E. coli RNase H is no less than 300,000 units per mg.
Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, (2nd Ed.), 8.64 – 8.65