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. 2022 Dec 23;9(1):27. doi: 10.3390/jof9010027

Figure 9.

Figure 9

Schematic illustration of the LAMP assay for detecting P. insidiosum. (a) In the initial step of the LAMP method, the F2 portion of the FIP primer anneals the F2c region of the DNA template to form a new DNA strand using the Bst DNA polymerase. The F3 primer anneals the F3c region of the target DNA to initiate sequence elongation, causing displacement of the FIP-linked strand. (b) The displaced strand forms a loop at the 5’ end and subsequently serves as a template for DNA amplification using the primers BIP and B3. (c) As a result, a loop is formed at both ends. The obtained double stem-loop DNA (a dumbbell-like structure) enters multiple amplification cycles for exponential production of the target DNA. The final LAMP product is a mixture of various sizes of stem-loop DNAs. (d) The LAMP products can be detected as a ladder pattern observed on an electrophoresis agarose gel. (e) Naked-eye observation of an amplification result under natural light after adding a fluorescent intercalating dye (SYBR Green I) shows the striking color change from orange to bright green, indicating a positive LAMP reaction. The diagram is adapted from the references [68,69] and the New England Biolabs website (https://international.neb.com, accessed on 6 November 2022).