DNA-based fish species identification protocol.

We have developed a fast, simple, and accurate DNA-based screening method to identify the fish species present in fresh and processed seafood samples. This versatile method employs PCR amplification of genomic DNA extracted from fish samples, followed by restriction fragment length polymorphism (RFLP) analysis to generate fragment patterns that can be resolved on the Agilent 2100 Bioanalyzer and matched to the correct species using RFLP pattern matching software. The fish identification method uses a simple, reliable, spin column- based protocol to isolate DNA from fish samples. The samples are treated with proteinase K to release the nucleic acids into solution. DNA is then isolated by suspending the sample in binding buffer and loading onto a micro- spin cup containing a silica- based fiber matrix. The nucleic acids in the sample bind to the fiber matrix. The immobilized nucleic acids are washed to remove contaminants, and total DNA is recovered in a final volume of 100 mul. The isolated DNA is ready for PCR amplification with the provided primers that bind to sequences found in all fish genomes. The PCR products are then digested with three different restriction enzymes and resolved on the Agilent 2100 Bioanalyzer. The fragment lengths produced in the digestion reactions can be used to determine the species of fish from which the DNA sample was prepared, using the RFLP pattern matching software containing a database of experimentally- derived RFLP patterns from commercially relevant fish species.

The effects of subgingival calculus on the clinical outcomes of locally-delivered controlled-release doxycycline compared to scaling and root planing.

BACKGROUND/AIM: The effect of subgingival calculus on the clinical outcomes of the local delivery of antimicrobials is unknown. This study examines the clinical outcomes of treatment with locally delivered controlled-release doxycycline (DH) or scaling and root planing (SRP) in subsets of adult periodontitis patients with known baseline levels of subgingival calculus. METHODS: The data examined were obtained from 393 patients who participated in 2 multi-center trials. All patients had baseline subgingival calculus levels assessed and were then treated at baseline and month 4 with either DH or SRP. Clinical attachment levels (CAL), pocket depth (PD) and bleeding on probing (BOP) were assessed at baseline and months 1, 2, 4, 5, 6, 8 and 9. RESULTS: Treatment with either DH or SRP resulted in significant statistical and clinical improvements in CAL, PD and BOP. These clinical outcomes were equivalent regardless of the extent of subgingival calculus present at baseline. CONCLUSIONS: The results indicate that the primary clinical effects of these therapies are the result of a disruption and reduction of the subgingival plaque and not the effect of the removal of subgingival calculus and contaminated cementum.