法医学研究ジャーナル

In order to perform human identification through DNA analysis, bones and teeth are considered the samples with the highest success rates. Regarding the protocols used to process these samples, they are also usually submitted to the same treatment, which consists of sample powdering. However, there are tissues in a tooth that have low DNA content, and thus, should not be included in DNA extraction. Recent studies have reported that sampling of dental cementum could optimize DNA typing from teeth due to its high cellularity and resilience. The objective of this article is to present an improved protocol for DNA extraction from teeth that targets cementum and its use in two real cases. After a 24-hour decalcification step, cementum was sampled and submitted to a simple DNA extraction protocol based on a commercial kit and automated platform. Real-time PCR (Polymerase Chain Reaction) quantification, PCR amplification of 27 DNA markers and capillary electrophoresis were performed. For both real cases, a different sample type had yielded a full DNA profile using a different and validated protocol. Quantification showed expected concentration and integrity of DNA in both tooth samples. After genotyping, all samples presented full and concordant profiles. This pilot study demonstrated that the presented protocol, specifically designed for teeth, was able to obtain full DNA profiles from a burnt and a skeletonized human body.

After the onset of the Syrian war in 2011, the Islamic State of Iraq and Syria (ISIS) have invaded a significant area of the country including a rural area at the north-eastern Lebanese-Syrian borders. Since then, there were several trials of invasion into the Lebanese territories until August 2014 when the north-eastern Lebanese town of Arsal was briefly invaded. During this invasion, 18 Lebanese Army soldiers were held hostage by ISIS and kidnapped out of the country into a no-mans-land at the Lebanese-Syrian borders.

In parallel, since the onset of the Syrian war, thousands of Syrians have been kidnapped and reported missing. Many of these crimes were especially committed against ethnic and religious minorities such as the Syrian Yazidi and Syrian Christians (Figure 1). These human rights violations are occurring on daily basis and led to thousands of mass graves, each having its characteristics, its difficulties and its mysteries that increase with the continuing war and the increasing number of missing individuals. However, the primary focus is to dedicate all the available means to determine the identity of each victim in order to return the remains to their families, in accordance with the Geneva conventions.

Touch or trace DNA analysis has become an important routine of the forensic laboratory workload and a useful tool for investigators. Most samples, such as touch DNA, are collected using cotton swabs and choosing the right collection technique when using a cotton swab can improve DNA recovery from the surfaces. Therefore, this paper investigates three recovery techniques commonly used with cotton swabs and validate different conditions on the collected swabs such as drying prior freezing or direct freezing to see how they affect the amount of DNA recovered. The results show that there is a significant difference between the three recovery techniques used to recover touch DNA with cotton swab (F_2,21 = 39.504, p<0.001), similarly with the cotton swab tested conditions prior extraction (F_2,21 = 68.328, p<0.001).

Background: Many molecular biology experiments start with DNA extraction. The quality and the quantity of the extracted DNA are very important for downstream analyses. Therefore, the DNA extraction process is very important and efforts should be aimed at constant improvement of the available protocols.

Objective: The first objective of this study is to compare various lysing agents that use Sodium Dodecyl Sulphate (SDS) in DNA extraction process. The second objective is to report detailed protocol for the extraction of DNA from frozen animal sample.

Methods: The efficiencies of four different lysing reagents involving SDS and a commercial lysing agent were compared. The reagents tested were ethylenediaminetetraacetic acid (EDTA), Sodium Chloride-Tris-EDTA (STE), Sodium chloride (NaCl) and tris acid (Tris). The four lysing agents investigated were SDS+EDTA+NaCl (SEN), SDS+EDTA+Tris (SET), SDS+STE+NaCl (SSN) and SDS+STE+Tris (STT). In addition, Qiagen (Q) lysing buffer was also included. The experiments were conducted in duplicates. The five lysing reagents were used to extract DNA from homogenized muscles and bones from the sternum of juvenile Japanese giant flying squirrel that had been frozen for four years.

Results: With the protocol presented here, gel electrophoresis, nanospec, bioanalyzer and PCR showed that all the five lysing reagents were able to extract DNA from the homogenized muscles and sternum bones of frozen rodent species. Large DNA molecules could be retrieved from all the lysing reagents. However, A260/A280 shows that SSN and STT had the purest extracted DNA molecules.

Conclusion: We present a protocol that successfully extracted DNA with good integrity from frozen animal sample. The comparison of five lysing reagents showed that although all the reagents successfully extracted DNA, the purest extracted DNA came from SDS+STE+NaCl (SSN) and SDS+STE+Tris (STT). Interestingly, DNA from all the reagents was successfully amplified with PCR suggesting that the impurities may not be of significant impacts to the downstream analyses.