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RNase H, E.coli

#EN0201 Supplied with: 10X Reaction Buffer	100 u (5 u/µl) 1 ml
#EN0202 Supplied with: 10X Reaction Buffer	500 u (5 u/µl) 1 ml
Related Documents (in pdf, ~40-60 KB): Certificate of Analysis: #EN0201, #EN0202 MSDS (English) MSDS (English-USA) MSDS (German)

Description
The RNase H, E.coli specifically degrades only the RNA strand in RNA-DNA hybrids. It does not hydrolyze the phosphodiester bonds within single-stranded and double-stranded DNA and RNA.

Source
E.coli MRE-600 cells.

Molecular Weight
18.4 kDa monomer.

Applications

• Removal of mRNA prior to the synthesis of the second strand of cDNA (1), see protocol.
• Removal of the poly(A) sequences of mRNA after hybridization with oligo(dT) (2).
• Site-specific cleavage of RNA (3).
• Studies of in vitro polyadenylation reaction products (4).

Quality Control
The absence of endodeoxyribonucleases, exodeoxyribonucleases and ribonucleases confirmed by appropriate quality tests.

Concentration
5 u/µl

Definition of Activity Unit
One unit of the enzyme catalyzes the formation of 1 nmol of acid soluble products in 20 min at 37°C.
Enzyme activity is assayed in the following mixture: 20 mM Tris-HCl (pH 7.8), 40 mM KCl, 8 mM MgCl₂, 1 mM DTT, 24 µM [³H]-poly(A)·poly(dT), 0.03 mg/ml BSA, 4% (v/v) glycerol.

Storage Buffer
The enzyme is supplied in: 25 mM HEPES-KOH (pH 8.0), 50 mM KCl, 1 mM EDTA, 1 mM DTT, 0.1 mg/ml BSA and 50% (v/v) glycerol.

10X Reaction Buffer
200 mM Tris-HCl (pH 7.8), 400 mM KCl, 80 mM MgCl₂, 10 mM DTT.

Inhibition and Inactivation

• Inhibitors: metal chelators, SH-blocking reagents.
• Inactivated by heating at 65°C for 10 min.

Related Products

M-MuLV Reverse Transcriptase RevertAid™ M-MuLV Reverse Transcriptase RevertAid™ H Minus M-MuLV Reverse Transcriptase DNA Polymerase I, E.coli Hot Start Taq DNA Polymerase TrueStart™ Taq DNA Polymerase Pfu DNA Polymerase Taq DNA Polymerase (native) Taq DNA Polymerase (recombinant) T4 DNA Ligase First Strand cDNA Synthesis Kit RevertAid™ First Strand cDNA Synthesis Kit RevertAid™ H Minus First Strand cDNA Synthesis Kit 0.5 M EDTA, pH 8.0 DEPC-treated Water

References

1. Gubler, U., Hoffman, B.J., A simple and very efficient method for generating cDNA libraries, Gene, 25, 263-269, 1983.
2. Davis, R., et al., Tandemly repeated exons encode 81-base repeats in multiple, developmentally regulated Schistosoma mansoni transcripts, Mol. Cell Biol., 8, 4745-4755, 1988.
3. Donis-Keller, H., Site specific enzymatic cleavage of RNA, Nucleic Acids Res., 7, 179-192, 1979.
4. Goodwin, E.C., Rottman, F.M., The use of RNase H and poly(A) junction oligonucleotides in the analysis of in vitro polyadenylation reaction products, Nucleic Acids Res., 20, 916, 1992.

Updated kovo 18, 2008 09:58