Ribonuclease H (RNase H)

Description

• General description: Nonspecific endoribonuclease that specifically cleaves RNA in RNA:DNA hybrids. A minimum of four continuous base pairs (RNA:DNA) is required for activity. RNase H cleaves RNA to release 5′-oligoribonucleotides.Source: E. coli H560 pol A1Storage Buffer: 25 mM Tris-HCl, 50 mM KCl, 1 mM dithiothreitol, 0.1 mM EDTA, 50% glycerol (v/v), pH 8.0 (+4°C)Volume Activity: 1 x 103 U/ml assayed according to Hillenbrand & Staudenbauer.
• Application: Ribonuclease H (RNase H) has been used for:In vivo RNA-primed initiation of DNA synthesisElimination of mRNA during second-strand cDNA synthesis[4]Site-specific cleavage of RNADetection of RNA:DNA regions in double-stranded DNA of natural origin[5]Removal of poly (A) sequences of mRNA if oligo (dT) is presentRNA extraction and quantitative reverse transcriptase polymerase chain reaction (RT-PCR)[6]
• Biochem/physiol Actions: Ribonuclease H (RNase H) specifically cleaves RNA in RNA:DNA hybrids.[1] A minimum of four continuous base pairs (RNA:DNA) is required for activity. RNase H cleaves RNA to release 5′-oligoribonucleotides. RNase H is associated with nucleic acid immunity.[1] Using RNase H for degrading mRNA results in 80% depletion of mRNA and protein expression. RNase H recognizes the start codon and the 3′ and 5′ untranslated regions.[2] This enzyme participates in DNA replication.[3]
• Features and Benefits: Eliminate potential sources of PCR errors.Increase accessibility of primers during subsequent PCR.
• Quality: Absence of endonucleases, nicking activities, and ribonucleases.
• Unit Definition: RNase H is assayed according to Hillenbrand and Staudenbauer. One unit of RNase H is the amount of enzyme which produces 1 nmol acid-soluble ribonucleotides from[3H] poly(A) x poly(dT) in 20 minutes at +37 °C under the stated assay conditions.Volume Activity: Approximately 1 U/μl
• Preparation Note: Activator: The enzyme has its maximal activity in presence of SH-reagents
• Storage and Stability: Store at -15–-25 °C. (unopened)
• Other Notes: For life science research only. Not for use in diagnostic procedures. Using RNase H after the cDNA synthesis step can increase the sensitivity of a two-step RT-PCR assay.