Application Guide - QIAxcel Advanced

QIAxcel Advanced Application Guide 10/2016 105 similar results. To assess utility of the method for processed commercial foodsamples, salmon spinach lasagna was digested with 4 enzymes, of which 1 enzyme was added for result confirmation (data not shown). Three different salmon species were identified in the lasagna, S. salar, Oncorhynchus gorbuscha and O. keta (Figure 2). The AluI digest was uninformative as it gave rise to the same band pattern for all 3 species, however, fragments from HaeIII, HinfI and DdeI digestion enabled discrimination of S. salar from O. keta and O. gorbusha. For example, the HaeIII restriction profile consisted of 4 bands, each coming from the band patterns of different salmons. The band at 41 bp was a fragment common to all three species, the bands at 109 bp and 309 bp came from S. salar, and the band at 419 bp came from O. keta/O. gorbusha. The two Oncorhynchus species were then identified using a fifth enzyme (data not shown). The PCR-RFLP method proved successful in fish species identification. Each fish species had a unique profile when using the set of enzymes described above. Correlation between theoretical and observed data was good for both fresh crude and frozen fish (data no shown). Moreover, the method enabled discrimination of mixtures containing up to 3 species. During the validation, we observed that some fish, like Theragra chalcogramma, had a polymorphic profile or point mutations. Such samples may give rise to false negatives or false positives, which must be taken into consideration for interpretation. The method also works well for several processed foods, such as salmon spinach lasagna, crab sticks or salmon parmentier, as long as the extracted DNA is not overly degraded. Only severly processed foods (e.g., canned rillettes) contain excessively degraded DNA that cannot be analyzed. Conclusions • The tested fish species generate unique digestion profiles and can be readily identified based on comparison to known profiles in a database. Up to 3 different species can be identified in mixtures. • Commercial samples of varying processing degree can be analyzed, as long as the DNA has not been overly degraded. • The QIAxcel Advanced facilitates the identification of fish species based on PCR-RFLP and provides results in less than 8 h. • Using the QIAxcel Advanced, the method is inexpensive and reliable, making it a good candidate for routine use in fish species identification. Fish species Theoretical band sizes (bp) Observed band sizes (bp) Differences S. scombrus 56 201 214 55 204 221 –1 +3 +7 M. merluccius 464 466 +2 S. salar 198 266 204 273 +6 +7 Table 3. Comparison of theoretical and observed band sizes arising from HinfI restriction digests M. merluccius 464466 +2

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