Ion force fast

DNA from the blank samples (10 replicates) and the spiked samples (10 replicates) for each matrix was extracted using ION Force FAST (Generon, San Prospero, MO, Italy). Briefly, 20 mL of solution A were added to a bottle containing 5 g of sample and incubated for 1 h at 85 °C in a water bath or in a thermomixer. After centrifugation for 10 min at 10,000 rpm, the aqueous supernatant was transferred to a 50-mL bottle, and an equal amount of buffer E was added. The samples were vortexed and centrifuged for 10 min at 10,000 rpm. A set of the subnatants were added with 7.5 mL of buffer T and 20 µL of glacial acetic acid. After filtering the samples in 12-mL syringes and 0.45 micron filters (this step was omitted for the yogurt matrix), the columns on the vacuum chamber were assembled, and the vacuum pump was operated for column purification. Two successive washes were then performed using 1 mL of buffer P. The column was centrifuged for 5 min at 7000 rpm to remove ethanol residues. DNA was eluted with 100 µL of buffer D after centrifugation at 500 rpm for 30 s and at 14,000 rpm for 5 min.

DNA extraction was performed on 10 test portions (replicates) from true negative blank samples and all spiked samples according to the commercial ION Force FAST (Generon, San Prospero, MO, Italy) kit. All protocol procedures were performed according to previously described protocols [23 (link)] and the manufacturer’s instructions.

In order to compare the performance of two different extraction kits, DNA extraction from each sample was carried out by means of QIAamp DNA Mini Kit (Qiagen, Milano, Italy) and Ion Force Fast (Generon, San Prospero, Modena, Italy). All extractions were carried out according to the manufacturers’ instructions.