FINDeM: A CRISPR-based, molecular method for rapid, inexpensive, and
field-deployable organism detection.
Abstract
The field of ecology has undergone a molecular revolution, with
researchers increasingly relying on DNA-based methods for organism
detection. Unfortunately, these techniques often require expensive
equipment, dedicated laboratory spaces, and specialized training in
molecular and computational techniques; limitations effectively
excluding field researchers, underfunded programs, and citizen
scientists from contributing to cutting-edge science. It is for these
reasons that we have designed a simplified, inexpensive method for
field-based molecular organism detection – FINDeM (Field-deployable
Isothermal Nucleotide-based Detection Method). In this approach, DNA is
extracted using chemical cell lysis and a cellulose filter disc,
followed by two body-heat inducible reactions – recombinase polymerase
amplification and a CRISPR-cas12a fluorescent reporter assay – to
amplify and detect target DNA, respectively. Here, we demonstrate FINDeM
in detecting Batrachochytrium dendrobatidis, the causative agent of
amphibian chytridiomycosis, and show that this approach can identify
single-digit DNA copies from epidermal swabs in under one hour using
low-cost supplies and field-friendly equipment.