What is the focus of your laboratory?
The general focus of my laboratory is forensic molecular genetics. We are trying to enhance the ability of the forensic scientist to extract information from biological evidence left at the crime scene, and to obtain probative information for utilization in court.
Our research group is currently approximately 12 people, consisting of Ph.D. students, master students, and staff. Embedded in the University of Central Florida, we are in a lucky environment with the opportunity to gather students from the UCF biomolecular science and forensic science programs.
What is your scientific background? Were you always doing forensic research?
As for my scientific education, I graduated with a Bachelor of Science Honors Degree in biochemistry in the 1970s and later with a Master of Forensic Science. In between I was a police officer in Great Britain and Hong Kong. Switching from the street to the laboratory — using brain instead of brawn so to speak — I became a forensic geneticist in the UK, later in Hong Kong again, and then in New York. During my lab tenure in Long Island, NY, I received a Ph.D. in genetics from the State University of New York at Stony Brook.
In best describing my work today, I would say it is conducting state-of-the-art forensic research while trying to help educate the next generation of forensic scientists. For that, I bring to the table a lot of practical experience from my time as police officer and forensic analyst, and I’m trying to apply the insights gained in order to advance methods for forensic investigation.
What types of projects and studies are underway?
Our work can be grouped in three main areas. Primarily we are conducting basic research. However we also have projects that test and evaluate new methods and projects that provide operational support to the community and to forensic laboratories. As an example of the latter we are managing and maintaining here at NCFS the US Y-STR Haplotype Reference Database. On the more fundamental research side, we focus on Y-chromosome analysis, body fluid identification, the area of physical characteristics, low-copy-number analysis, and damage and repair of DNA.
Y-chromosome analysis is important for sexual assault investigations and, in the future for helping predict a male individual’s biogeographical ancestry. We have developed a number of Y-STR and Y-SNP multiplex systems to accomplish these goals.
We have spent a lot of time developing mRNA profiling methods for the identification of body fluids and tissue. Our goal is to substitute molecular genetics techniques for standard immunological methods that are slow, not very specific, and consume precious sample. In practice, we use methods that take advantage of our ability to simultaneously co-isolate DNA and RNA portion from the same sample.
Another work focus is the area of physical characteristics. A STR-based DNA profile itself does not in itself tell us anything about the physical attributes of the person depositing the stain. It can only be used to assess identity or disparity of biological case evidence to an established reference, a suspect, or a database. But DNA can be further interrogated and serve as a ‘genetic eyewitness’, so to speak: sex, height, weight, and stature; hair, eye and skin color; as well as age and ethnic background can, in principle, be determined on a genetic level. DNA (or even RNA-) based physical characteristics data will one day complement the DNA profile or deliver crucial information in the absence of a database hit. DNA-encoded age information is particularly fascinating. Imagine a terrorist case where DNA traces of a senior person would point to the bomb engineer or ”mastermind” while the DNA of a younger person could be attributed to the assailant.
Low-copy-number analysis is another one of our focus areas. Our interest lies at the level of a single cell or even beyond. We term it “smart” single-cell analysis, as one is able to avoid the analysis of mixtures by looking at recovered single cells, for instance, by means of laser capture microdissection. Currently comparatively large samples are collected from the crime scene as touch DNA samples that are prone to contamination and often comprise mixed samples.
Last but not least, we are interested in the principles of damage and repair in DNA evidence. We are trying to assess the quality of DNA, its state of degradation, and we are experimenting with biochemical in vitro reconstitution of DNA repair activities.
What are the main techniques utilized in the lab?
We use the standard molecular biology equipment needed to perform DNA amplification (PCR, quantitative PCR, RT-PCR, quantitative RT-PCR) and forensic DNA analysis (capillary electrophoresis). We also employ a variety of other standard molecular biology methods such as cloning and sequencing. Low-copy-number techniques are being made use of, including whole genome amplification. We are also using Pyrosequencing and laser-capture microdissection technologies. We are beginning to use mass spectroscopy (MS) in combination with liquid chromatography (LC) methods for bioanalyte separations.
How important is the preanalytical part of the forensic analysis process — sample collection, stabilization, and extraction — for the overall result?
Without an appropriate preanalytical process, one’s analytical results are meaningless in a forensic context. One has to ensure that the material one is looking at in the laboratory is indeed the material purportedly left at the crime scene. The integrity of the evidence has to be maintained using all means possible. It must not be cross-contaminated and the evidence must not be permitted to degrade or be damaged any further.
Evidence must be properly recorded,collected, and preserved, in as few steps as possible. The forensic process in that respect is more demanding than a clinical process. The forensic process deals with minuscule amounts of DNA — nanogram or even sub-nanogram quantities — while by comparison the clinical sampling process mostly results in microgram yields .
One type of information is not yet being deduced from the crime scene by molecular means: (point of) time. How do you see that changing?
At a crime scene one has to distinguish between signal and noise: information that is relevant to the crime (‘signal’) and information that is not ("noise"). A person could have had legitimate access to the scene prior to the crime — which would be an alibi. Now, being able to determine when biological material was deposited at the scene would help to determine the legitimacy of a suspect’s alibi.
Degradation to macromolecules such as DNA, RNA, or proteins in biological stains may serve as a molecular clock . Alternatively, one can look at the accumulation of monomeric or low-molecular-weight molecules. For example with proteins, the number of enzymes still present and their associated enzyme activity changes over time in dried stains. Any such predictor of time since deposition must be looked at in the context of multiple variables, with heat and humidity being the most important ones. The goal is to determine if hours, days, week, months, or years have passed since the crime was committed. It will be a significant challenge to obtain higher resolution than this, though.
You run a research laboratory. How do you get access to “real life” forensic samples?
We are doing this in two ways: by creation of mock up samples and by partnerships.
We have a profound understanding of what forensic evidence looks like, allowing us to design “fake” samples. For instance, we produce stains on clothes and expose them to various environmental conditions, and we have volunteers donating post coital samples. We also partner with a number of operational crime labs to obtain a variety of bona fide casework samples.
One of the technologies you are investigating is whole genome amplification (WGA). How could WGA help the forensic investigator in the future? Will it become a routine application?
Whole genome amplification is one way of dramatically increasing the sensitivity of analysis of minute forensic samples while still permitting the production of reproducible and consistent genetic information. Current nuclear DNA techniques have sensitivity levels in the range of hundreds of picograms. We want to be able to extract accurate genetic information when the sample yields less than 100 pg, or even down to the cell level on a routine basis. Our hope is that WGA will help to increase sensitivity without some of the disadvantages of other highly sensitive low copy number methods, such as electrophoretic artifacts.
What role do you see for automation (for extraction/detection) etc. in the forensic lab now and in the future?
What are your most important criteria for automating a procedure such as, for example, sample extraction?
Automation is an absolute requirement for us for the future. We need timely analysis. When people talk about the quality of the forensic analysis, they often think of the “error proof” aspect of automation. But timeliness is equally important in many circumstances. We have a limited number of staff, and human resources are costly. Automation frees up the scientist to look at the case in a holistic manner and focus more on the important aspects — the up front side (i.e., what has or should be analyzed and for what?) and the interpretation of the analytical results — instead of the analytical process itself.
As for automation criteria, we require accuracy and reliability, with at least the same accuracy and reliability as the established manual method. Evidence cross-contamination studies of robotic systems are important, and the instruments must be easily maintainable, along with swift and good technical support.
What are the biggest challenges for your laboratory?
I think our biggest challenge is how to transfer technologies and methods that we develop into operational use. Because of extensive validation requirements that operational labs have to follow prior to implementation of a new technology and the level of sophistication of some of the new methods, it is difficult to seamlessly transfer such technologies from academic research into operations.
Secondly, forensic research funding: the classical academic funding sources (e.g. National Institutes of Health and the National Science Foundation) are unavailable to forensic researchers.
If you were to envision molecular forensics 10 years from now, which technologies would be around, what kind of data and information would be derived?
Forensic labs will be fully automated and the associated personnel structure will be different: We’ll see fewer people involved in the analytical pipeline process itself. More people will be working at a higher level doing data interpretation and “piecing things together”. In some ways it will look physically like the clinical biochemistry labs of today with bank upon bank of automated equipment. Probative genetic Information will be acquired very rapidly. This will permit the rapid, wireless, remote interrogation of various DNA databases. DNA profiles will be obtained within an hour of sample acquisition (or less) and with significantly less human intervention.
As for technologies: miniaturization/lab-on-a-chip and nanotechnologies will come into play. Will we still be using STR markers? Most forensic scientists believe that we will, but this not entirely clear.
As for the types of information that will be gathered from a deposited body fluid stain, one should be able to get a profile or unique DNA identifier, the nature of the body fluid or tissue source from which the profile was obtained , and “genetic eyewitness information” (physical characteristics). This may even include information about the facial features or even the behavioral characteristics of the donor. I would not be surprised if everybody’s DNA profile is encapsulated in population-wide databases that were obtained shortly after birth.
