Removal of Contaminating Genomic DNA in QRT-PCR using a Shrimp Nuclease

DNA contamination can often occur in quantitative reverse transcription – polymerase chain reactions (QRT-PCR), and should be removed in order to avoid false positive results.  DNase I is commonly used for removing DNA contamination, but this has a relatively long and harsh protocol which introduces an extra step between the isolation of RNA and the QRT-PCR reaction itself, as well as increasing the risk of RNA degradation due to the harsh inactivation conditions.

An endonuclease from arctic shrimp Pandalus borealis has properties that make it useful for the removal of contaminating DNA. The endonuclease activity of the enzyme is specific to double stranded DNA, which therefore allows the enzyme to be added directly into the reverse transcription step.  Unlike DNase I, the shrimp nuclease is easily inactivated at high temperatures, such as those used for the RT deactivation / hot start incubation step of a QRT-PCR reaction, and therefore can be used to selectively degrade double stranded DNA, leaving single stranded DNA and RNA intact.