Using CRISPR/Cas12a to detect new coronavirus (COVID-19:SARS-CoV-2): Sensitization effect adding Mn2+

CRISPR/Cas9 won the 2020 Nobel Prize in Chemistry about a week ago.
We have found several papers that used this technique as a high sensitive detection method for the new coronavirus. The following paper reports that detection using CRISPR/Cas12a could be further 13 times more sensitive by adding Mn2+.

A brief description of the protocol is as follows:

  • Extract an RNA sample from a sample.
  • Make cDNA using reverse transcription, and amplify by PCR (dsDNA is made).
  • Pre-design crisper RNA (crRNA) using RNA sequences specific to the E protein of the new coronavirus.
  • Add Cas12a enzyme and crRNA to the dsDNA sample, the new coronavirus-specific sequence is cut out when it is in the sample dsDNA.
  • Thus, since the inhibitory interaction between the fluorescent probe and the quencher that have been incorporated in the PCR process is lost, the fluorescence comes out by applying the excitation light.

Of course, this fluorescence is read by a detector, but it can be confirmed by the eyes.
The addition of Mn2+ to this system increased the sensitivity by 13 times.