FastDigest Eco72I

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	Briefly, 5 µL of the amplification products was digested using the restriction enzyme PmaCI (FastDigest Eco72I, ThemoFisher) for 15 min at 37 ℃ followed by agarose gel electrophoresis of the restriction fragments on a 2% gel stained with ethidium bromide. The fragments were visualized in a UV transilluminator and photographed.

Protocol tips
Briefly, 5 µL of the amplification products was digested using the restriction enzyme PmaCI (FastDigest Eco72I, ThemoFisher) for 15 min at 37 ℃ followed by agarose gel electrophoresis of the restriction fragments on a 2% gel stained with ethidium bromide. The fragments were visualized in a UV transilluminator and photographed.

Publication protocol

The 16S rRNA gene was amplified from the genomic DNA samples of 13 P. salmonis strains and 57 fish tissues (field samples) as described in Section 4.5. Initially, the presence and purity of P. salmonis in the field samples were verified as described in Mandakovic et al. [18]. Briefly, 5 µL of the amplification products was digested using the restriction enzyme PmaCI (FastDigest Eco72I, ThemoFisher) for 15 min at 37 ℃ followed by agarose gel electrophoresis of the restriction fragments on a 2% gel stained with ethidium bromide. The fragments were visualized in a UV transilluminator and photographed. Then, the PCR amplification products from the P. salmonis strains and field samples were subjected to a XapI enzymatic digestion. The reactions were carried out in 15 μL volumes, which contained 5 μL of the PCR product, 1.5 μL of enzyme buffer, 1 U of each enzyme and 8 μL of nuclease-free water, for 30 min at 37 °C. The restriction fragments were visualized by electrophoresis in 2% agarose gels stained with 0.5 µg/mL of ethidium bromide for 30 min at room temperature.

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Manufacturer protocol

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