NEW YORK (GenomeWeb) – A consortium established by the European Commission in 2012 is working to develop new methods and establish standards for DNA profiling by harnessing advances in next-generation sequencing and examining the ethical, legal, and social implications of increasingly sensitive forensic methods.
The European Forensic Genetic Network of Excellence (EUROFORGEN-NoE) recently described the validation of a SNP-based NGS panel to identify ancestry of a DNA sample in the journal Forensic Science International: Genetics.
Peter Schneider, project coordinator of EUROFORGEN-NoE and a professor at the University of Cologne's Institute of Legal Medicine in Germany, told GenomeWeb that the EC funded the group in 2012 for five years, after which the goal is to become self sustaining. Currently, 16 partners from nine different European countries make up the consortium.
The project is divided into five different "work packages," Schneider said. One is the organizational component that facilitates communication between the consortium members. About half of the funding supports research projects, which are divided into two more components — one that focuses on developing new DNA profiling methods, and another that focuses on specific applications such as prediction of ancestry and physical characteristics. Fourth, there is an ethical, legal, and social component that will develop ethical guidelines and look at the risks and privacy issues created by more advanced NGS-based DNA profiling methods. Finally, a fifth group is related to education and aims to develop a postgraduate curriculum for forensic genetics.
When the group originally formed in 2012, Schneider said that NGS technology was not yet at the point where it was feasible to be used for forensic applications due to the high cost and the fact that the instruments required so much input DNA.
But now, he said, much of the group's work is focused on developing NGS-based methods. The technology enables "much greater sensitivity and complexity — for instance, from one sample you can identify ancestry, and make predictions about the individual's hair and eye color," he said. It is also better at unraveling samples that contain DNA from multiple individuals and multiple sample types.
In addition, researchers are testing approaches to use NGS to make age predictions by looking at DNA methylation, Schneider said.
Another new area of forensic research that the EUROGEN-NoE consortium is just starting to investigate is in analyzing RNA or microRNA, which can enable scientists to identify the tissue of origin of a sample. This could be particularly useful for sexual assault cases where there could be genetic material from blood, semen, and vaginal fluid. By looking at RNA, though, scientists can "distinguish between saliva and semen, for instance, which helps to get a better idea of how that sample originated," Schneider said.
In their recent study, the group published a validation of an ancestry SNP panel. Schneider said the panel includes 128 SNPs that could distinguish the five major populations — European, Asian, Oceanian, American Indian, and African. In addition, he said, the group added SNPs that could distinguish between admixed populations. For example, individuals from Southeast Asia typically have both European and Asian ancestry.
They designed the panel on Thermo Fisher Scientific's Ion Torrent PGM using the AmpliSeq chemistry. Schneider said one important facet was being able to get good results from small amounts of input DNA. The group was able to get a full profile using as little as 100 picograms of DNA, he said, which is about the equivalent of 15 cells.
The researchers tested the panel on diluted samples, degraded DNA, and DNA mixtures. In addition, they tested the panel at five different laboratories to verify that it was reproducible.
One issue the researchers identified involved the location of the SNPs. From the original set of 128 SNPs, three "presented insurmountable problems for primer design," the authors wrote, due to being too close to repetitive regions. In addition, two of the three SNPs were also too close to homopolymeric regions, which created issues with sequencing, even if the researchers were able to successfully design primers. They were able to successfully replace two of the SNPs, but the third was in a region that had to be abandoned completely.
Aside from those problems, they found that the panel performed well across laboratories. In addition, while the panel was able to differentiate between subpopulations, the authors noted that a next step would be to improve upon this capability. For instance, in analysing Somali populations they found that the panel tended to underestimate the South Asian genetic contribution. So, for forensic laboratories with a significant proportion of South Asian and Middle East populations in their region, it would be worthwhile to add additional SNPs, they wrote.
"Ancestry-informative markers can potentially add useful information to the challenging task of deconvoluting mixed DNA patterns by allowing the inference of the ancestry of components in simple mixtures," the author wrote. Such a panel could be used from crime scene cases where investigators obtain a good DNA profile, but have no matches in the database. An ancestry-informative panel could help narrow down the range of suspects.
Schneider said that although a lot of research is being done on using NGS in forensics, it will be a while before such techniques are used regularly, particularly for case work.
"For a large number of cases, you don't need all that complex information," he said, because often once a DNA profile is generated there is a match in the database. As such, "for the time being, CE sequencing will be the standard."
Other forensic researchers who have been testing NGS technology have come to similar conclusions. For instance, researchers testing Thermo Fisher's new forensic sequencing panels have said that they are excellent research tools but that it would take time before they are used in casework. In the US, Thermo Fisher is looking to validate its panels through the Scientific Working Group on DNA Analysis Methods (SWGDAM), and Illumina, which also markets a forensic specific sequencing platform and panels, MiSeq FGx, has brought its system through SWGDAM validation.
These efforts to validate the technology, including by the EUROFORGEN-NoE consortium, are necessary steps.
Schneider anticipated that NGS-based panels would increasingly become adopted, first by specialized forensic labs for difficult cases. Before NGS can be used on routine casework though, cost and speed will both have to be reduced, and methods will have to be automated, simplified, and validated, he added.