A comparison between petrous bone and tooth, femur and tibia DNA analysis from degraded skeletal remains

Abstract

Skeletal remains are the only biological material that remains after long periods; however, environmental conditions such as temperature, humidity, and pH affect DNA preservation, turning skeletal remains into a challenging sample for DNA laboratories. Sample selection is a key factor, and femur and tooth have been traditionally recommended as the best substrate of genetic material. Recently, petrous bone (cochlear area) has been suggested as a better option due to its DNA yield. This research aims to evaluate the efficiency of petrous bone compared to other cranium samples (tooth) and postcranial long bones (femur and tibia). A total amount of 88 samples were selected from 38 different individuals. The samples were extracted by using an organic extraction protocol, DNA quantification by Quantifiler Trio kit and amplified with GlobalFiler kit. Results show that petrous bone outperforms other bone remains in quantification data, yielding 15–30 times more DNA than the others. DNA profile data presented likeness between petrous bone and tooth regarding detected alleles; however, the amount of DNA extracted in petrous bones allowed us to obtain more informative DNA profiles with superior quality. In conclusion, petrous bone or teeth sampling is recommended if DNA typing is going to be performed with environmentally degraded skeletal remains.

Evaluation of a New DNA Extraction Method on Challenging Bone Samples Recovered from a WWII Mass Grave

“…Overall, our results partially agree with previous data highlighting that metatarsals are ideal bone elements for genetic typing [ 21 ], even if a possible explanation for our finding could be ascribed to the peculiar burial conditions of the mass grave of the isle of Cres [ 3 , 14 , 44 , 45 , 46 , 47 ]. In addition, the low-quality genetic results obtained for molar teeth in our study do not support the positive outcome described for these elements recovered from some mass graves [ 14 , 38 , 42 ]. Again, as reported in several studies [ 3 , 12 , 14 , 44 , 45 , 46 , 47 ], environmental burial conditions (i.e., temperature and soil properties such as pH and ionic composition, hydrology, etc.)…”

“…As this method allowed no or very poor qPCR quantification values, we decided to increase the amount of starting bone powder to 0.5 g and set up a protocol based on the employment of the Maxwell ® FSC DNA IQ™ Casework Kit (Promega). The main advantage of this semi-automated DNA extraction protocol compared to other protocols, some of which are based on organic purification [ 3 , 14 , 31 , 42 , 43 ], is the decrease in the risk of human error and cross-contamination of the samples due to the automation of the DNA purification steps, especially by reducing physical manipulations of the specimens. The bone lysates, in fact, are transferred from 50 mL Falcon tubes into the cartridge after three simple and quick steps: (1) short centrifugation (in an Eppendorf tube) to pellet the undigested bone powder (if any); (2) transferring the sample to a second Eppendorf tube and mixing it with the buffer provided in the kit; (3) loading the sample into the prefilled cartridge.…”

“…Overall, the results of this study support that burial conditions play a fundamental role in biomolecule preservation, and no generalisation is allowed. Femurs, which usually provide satisfying results [ 3 , 14 , 38 , 42 ], yielded approximately ten-fold less DNA than other femurs found in a different WWII mass grave studied by us [ 33 ]. Similarly, teeth found in the mass grave of the isle of Cres yielded low levels of DNA.…”