Recycling of Polymerase Chain Reaction (PCR) kits

ABSTRACT During the outbreak of the SARS-CoV-2 pandemic, PCR (polymerase chain reaction) kits have been used as a common diagnosing method, with daily worldwide usage in the millions. It is well known that at the beginning of the pandemic there was a shortage of PCR kits. So far, the ecosystem of PCR kit is linear use, that is kits are produced, used one-time, and disposed in biolab wastes. Here we show that, to mitigate the risk of future shortages, it is possible to envision recyclable PCR kits, based on a more sustainable use of nucleic acid resources. A PCR kit is mainly composed of primers, nucleotides, and enzymes. In the case of a positive test, the free nucleotides are polymerized onto the primers to form longer DNA strands. Our approach depolymerizes such strands keeping the primers and regenerating the nucleotides, i . e ., returning the nucleic acid materials to the original state. The polymerized long DNA strands are hydrolyzed into nucleotides monophosphates that are then phosphorylated in triphosphates using a method that is a development of a recently published one. We used oligonucleotides with 3’-terminal phosphorothioate (PS) backbone modification as non-hydrolysable PCR primers, so to undergo the recycling process unchanged. We have successfully recycled both PCR primers (∼65% yield for 4-PS modification, and ∼40% yield for 2-PS modification) and nucleotides (∼75% yield). We demonstrate that the method allows for direct re-use of the PCR kits. We also show that the recycled primers can be isolated and then added to end point or quantitative PCR. This recycling approach provides a new path for circularly reusing PCR nucleic acids.