The adoption of next-generation sequencing (NGS) technology in the forensic industry is in its infancy, but its ability to garner more actionable information, particularly in compromised samples, is undeniable. Better described as massively parallel sequencing (MPS), the technology was introduced in the 21st century and has fundamentally revolutionized how biological systems could be studied with unprecedented throughput, scalability and speed. Important discoveries already have been made in many fields, in which, it is applied. For the forensics community in particular, MPS offers a highly complementary tool to Capillary Electrophoresis (CE) that is helping scientists in the field delve deeper into genetic information and find answers to very challenging questions.

CE sequencing and MPS are similar in that both methods rely on a DNA polymerase to drive incorporation of deoxyribonucleotide triphosphates into a DNA template strand in the sequential cycles of DNA synthesis. During CE sequencing, nucleotides are identified by fluorophore excitation, while nucleotide incorporation during MPS is measured by chemical- or fluoresence-based detection methods. The complementary nature of both systems lies in the combination of data generated by each.

Forensic scientists use CE to rapidly and cost-effectively test a validated set of short-tandem repeat (STR) markers by determining differences in DNA fragment lengths. MPS takes it a step further by, in addition to length information, providing STR sequence information and genotyping single nucleotide polymorphisms (SNPs) within the repeat marker sites. Also SNPs at other places within the genome can provide identity, ancestry and phenotypic information about the sample. While applying MPS to forensics may be perceived as taking a longer time to obtain a result, it can provide more genetic data from challenging or degraded samples and, on a per marker basis, in actuality is quite cost-competitive.

Mining for Clues from Unknown Human Remains

An example of these powerful technologies in action involves the case of unidentified human remains discovered in the small South Dakota town of Deadwood. Established in 1876, Deadwood was a primitive gold mining camp inhabited by thousands of pioneers, including historical figures such as James Butler “Wild Bill” Hickok, Seth Bullock, and Martha Jane “Calamity Jane” Cannary.

Little is left from the town’s lawless frontier days, but a recent study using CE and MPS technology has provided new insight into one former resident from that era who construction workers unearthed in 2012 while building a retaining wall near the city’s original cemetery site.

Using ancestry informative and phenotype markers, Y-STRs, and mitochondrial DNA analyses, scientists from the Institute of Applied Genetics at the University of North Texas Health Science Center were able to pull substantially more information from the unidentified, 140-year-old human skeleton than possible by traditional CE analyses. Prior to applying MPS, anthropological insights and a few lineage genetic markers were the only available points of reference for the former Deadwood resident. These techniques were able to determine that the person was a male and likely of European descent.

By sequencing the samples, however, the research team could now paint a more vivid picture of what this person once looked like. In addition to revealing his gender and ethnicity, the MPS data also suggested the man likely had light red hair and brown eyes. Remarkably, the team was able to generate the data and reach their conclusion in just a few days.


Indeed, MPS’s ability to unearth a wealth of data to supplement our field’s gold standard methods has significant potential to help solve modern-day crime as it moves from research to a validated technology. The tool has been proving its utility since its debut more than a decade ago, and there are many examples in which its ability to read data from highly degraded samples could have been instrumental to help solve cases, including a two-decades-old missing person investigation in particular.

Donna Williamson was a 19-year-old woman from North Richland Hills, Texas, who disappeared in August 1982. Her remains were found 11 years later on the side of a highway, but identity could not be established due to the degraded state of the bones and the limited technology available at the time. The physical examination only revealed the remains were likely female and provided approximate age and height. Unfortunately, in this case, human error may have delayed identification since it was later determined that investigators were comparing teeth taken from the right jaw to dental records of the left mandible. Williamson’s case would remain unsolved for another decade until DNA databasing in Texas came online.

By comparing DNA samples provided by Williamson’s family members with CE sequencing data recovered from her degraded samples, scientists were able to determine that there was a 95.21 percent probability that the unidentified remains belonged to Williamson. While not quite enough to positively identify her by DNA alone, investigators decided to re-examine dental records, which revealed the error in the original investigation. This cross-referencing effort helped positively identify Williamson’s remains. Using orthogonal methods for human identification may reduce such errors early on. If it had been available, MPS technology could have been that additional tool to help solve the mystery much earlier, and relatively quickly, with its ability to type more markers.

It is clear that MPS is moving the needle in every field it reaches. Similar to the critical advances MPS is affording science to better understand cancer, infectious disease, and even biodiversity, the technology has real potential to revolutionize public safety and the way we approach criminal investigations. Its use in compelling forensics research studies around the globe already is demonstrating how it could be applied in real-world cases. European countries like Denmark have validated the technology, and are currently using it for routine ancestry and human identity case work. The Netherlands also has implemented a similar MPS program to help solve crimes in that country, while the U.S. Armed Forces DNA Identification Laboratory has begun using MPS to help identify unknown remains from past wars. Time will tell how quickly MPS will be widely adopted as a validated tool for routine forensics in the United States, but the insightful research being generated today offers compelling reason for its use in crime labs everywhere.

Bruce Budowle, Ph.D. is director of the University of North Texas Health Science Center's Institute of Investigative Genetics (IAG) and Professor of the Department of Molecular and Medical Genetics. He is a renowned expert in forensic genetics and counterterrorism, primarily in identification of victims from mass disasters and microbial forensics. Prior to joining the UNT Health Science Center, Budowle spent 26 years as a scientist for the Federal Bureau of Investigation (FBI) Laboratory Division. He was a principal advisor in efforts to identify victims from the World Trade Center attack in 2001 and helped establish a mitochondrial DNA sequencing program to enable high-throughput sequencing of human remains.

Angie Ambers received her Ph.D. in molecular biology with an emphasis in forensic genetics and human identification. Her research has involved investigation of methods (e.g., whole genome amplification, DNA repair) for improving autosomal and Y-STR typing of degraded and low copy DNA from human skeletal remains and environmentally-damaged biological materials. She also holds master’s degrees in forensic genetics from the University of North Texas Health Science Center and in criminology from the University of Texas at Arlington. Prior to pursuing a doctorate, Dr. Ambers was lead DNA analyst and lab manager of UNT's DNA Sequencing Core Facility. Currently Dr. Ambers is a postdoctoral fellow at the IAG (specializing in skeletal remains analyses) and an adjunct professor at UNT, where she teaches Forensic Molecular Biology to students in the FEPAC-accredited forensic science program.