Optimizing DNA extraction from environmentally degraded bone samples for molecular identification of cetacean species

Molecular identification of cetacean bone remains can be hampered by DNA degradation and the presence of PCR inhibitors. Researchers have now developed an optimized DNA extraction protocol for environmentally exposed cetacean bones. This new method involves processing 100 mg of bone powder and subjecting it to a 24-hour digestion process using EDTA, N-lauroylsarcosine, and proteinase K. The sample is then purified using a modified silica-column technique.

The protocol was tested on nine environmentally degraded bone samples, representing eight different cetacean individuals. The resulting DNA concentrations varied between 7.3 and 57.1 ng/uL, with an average of 25.91-13.91 ng/uL.

Using conventional PCR, the mitochondrial cytochrome b gene was successfully amplified from all nine samples, and five samples (55.6%) produced sequences suitable for further analysis. BLASTn results identified Balaenoptera physalus as the closest match for all recovered sequences. A phylogenetic analysis further corroborated the association of these sequences with the B. physalus reference sequences.

These findings indicate that the proposed DNA extraction protocol offers a practical approach for recovering amplifiable and molecularly informative mitochondrial DNA from environmentally degraded cetacean bone material. This advancement enables molecular identification of cetacean species from challenging skeletal remains where DNA degradation and PCR inhibitors are prevalent.