A: RNase H is an endonuclease that specifically degrades the RNA strand of RNA:DNA hybrids while leaving the DNA strand intact. It does not act on single-stranded or double-stranded RNA or DNA.
A: RNase H selectively cleaves RNA that is hybridized to DNA. This activity is essential in workflows where RNA must be removed after serving as a template, such as during cDNA synthesis or nucleic acid amplification.
A: No. RNase H does not recognize specific nucleotide sequences. Cleavage occurs wherever RNA is hybridized to DNA.
A: No. RNase H only cleaves RNA when it is hybridized to DNA. Unhybridized RNA remains intact.
A: RNase H only cleaves RNA within RNA:DNA hybrids. In contrast, both RNase A and RNase I cleave single-stranded RNA.
A: Yes. RNase H requires divalent metal ions, typically Mg²⁺ or Mn²⁺, for catalytic activity. Chelating agents such as EDTA will inhibit the enzyme.
A: Yes. Incubation at 65°C for approximately 20 minutes effectively inactivates E. coli RNase H, depending on buffer conditions.
A: No. ScriptGuard™ RNase Inhibitor inhibits RNase A-family enzymes that target single-stranded RNA. RNase H belongs to a different enzyme family with distinct substrate specificity and is not affected.
A: After first-strand cDNA synthesis, RNase H partially degrades the RNA template, creating nicks and short RNA fragments. These fragments facilitate DNA polymerase-mediated synthesis of the second DNA strand.
A: Yes. DNA oligonucleotides can be designed to hybridize to specific RNA sequences, forming RNA:DNA hybrids. RNase H then cleaves the RNA at these targeted sites, enabling sequence-directed RNA cleavage.
A: In Nucleic Acid Sequenced Based Amplification (NASBA®), RNase H removes the RNA template following reverse transcription. This enables efficient DNA amplification and prevents RNA interference in downstream reactions.
A: Yes. Removing RNA after reverse transcription can reduce background interference and improve DNA amplification specificity and consistency.
