Epicentre"s® patented* Hybridase™ Thermostable RNase H specifically degrades only RNA in an DNA:RNA hybrid, without affecting DNA or unhybridised RNA. In contrast to E. coli RNase H, which is rapidly inactivated at 55°C, Hybridase RNase H is active at high temperatures. It has optimal activity above 65°C and can be used at temperatures up to 95°C. The thermostability of the enzyme permits it to be used at temperatures that give the highest hybridisation stringency for specific DNA:RNA heteroduplexes, maximising sensitivity and selectivity while minimising background due to nonspecific hybridisation.

Unit Definition

One unit results in the acid-solubilisation of 1nmole of 3H-polyadenylic acid in the presence of an equimolar concentration of polythymidylic acid in 20 minutes at 45°C in 50mM Tris-HCl, pH 7.5, 100mM NaCl, and 10mM MgCl2.Note: The unit assay is performed at 45°C because this is optimal for the Tm of poly(dT):poly(A). The optimal temperature for many applications may be considerably higher.

Storage Buffer

50% glycerol containing 50mMTris-HCl, pH 7.5, 0.1M NaCl, 1.0 mM DTT, 0.1mM EDTA, and 0.1% Triton® X-100.

Quality Control

Hybridase Thermostable RNase H is tested for RNA degradation in an RNA:DNA hybrid and for the absence of detectable DNA exo- or endonuclease, and non-RNase H RNase activities.

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Protocol for:Hybridase™ Thermostable RNase H

Due to the constant updating of the protocols by the manufacturer we have provided a direct link to Epicentre’s product page, where the latest protocol is available.

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Hybridase™ Thermostable RNase H Protocol

(catalogue numberH39100/ H39500)

Please note: all protocols off site are the responsibility of the products supplier

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References

1. Duck, P. et al. (1990) BioTechniques 9, 142.
2. Bekkaoui, F. et al. (1996) BioTechniques 20, 240.
3. Guatelli, J.C. et al. (1990) Proc. Natl. Acad. Sci. USA 87, 1874.
4. Fahy, E. et al. (1991) PCR Methods and Applications 1, 25.
5. Gutgsell, N. S. & Jain, C. (2010) Coordinated Regulation of 23S rRNA Maturation in Escherichia coli, J. Bacteriol. 192 , 1405-1409.
6. Laveder, P., et al. (2002) A two-step strategy for constructing specifically self-subtracted cDNA libraries, Nucleic Acids Res. 30 , e38-.
7. Parlow, M. H., et al. (1990) Enrichment of ubiquitinated histone H2A in a low salt extract of micrococcal nuclease-digested myotube nuclei, J. Biol. Chem. 265 , 7507-7512.

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Applications

• High-stringency hybrid selection.
• Diagnostic assays of specific target DNA sequences by isothermal probe amplification.^1,2
• Transcription-based amplification methods (e.g., NASBA® method).^3,4
• High-stringency mapping of mRNA structure.
• Applications that require specific hydrolysis of the RNA in a DNA:RNA hybrid.

If you cannot find the answer to your problem below then please contact us or telephone 01954 210 200