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Partial Profiles and Secondary Transfer of DNA

When an analyst is presented with an insufficient amount of DNA to obtain a full profile, should an individual be included in a mixture? Should we assume the remainder of the profile by resonable assumption or should we find further evidence/arguments. Secondary transfer of DNA is being studied due to potentially incarcerating the innocent.


In 1988 I obtained my Bachelor of Science Degree from Virginia Commonwealth University in Richmond, VA. My degree was issued by the School of Education whereby I obtained a total 32 credit hours in Chemistry, beyond the required 29 credit hours to be able to receive my accreditation to teach Chemistry and Math in the Commonwealth of Virginia. In 1989, I began employment with Commonwealth Laboratories in Richmond, Va., where I gained experience, knowledge, and training on laboratory methods, procedures, analysis, and report writing. In 1990, I began employment with Roche Analytics Laboratories (now LabCorp) where I gained further experience, knowledge, and training on laboratory methods, procedures, analysis, and report writing. The levels of training at both facilities included but were not limited to, chain of custody reports, sample preparation and digestion for analysis, analysis by wet chemistry/color analysis, infrared spectroscopy, liquid chromatography, gas chromatography, gas chromatography/mass spectroscopy, atomic absorption, inductively coupled plasma and inductively coupled plasma with ultrasonic nebulization. During my employment with these facilities, I obtained on the job training and gained certificates of completion for any formal instrument training I attended. Additionally, I was involved in new method development and assisted in research for lower levels of detection for some analytes. In 1995, I opened my own pharmaceutical laboratory designed for research and development for new pharmaceutical products. I have filed 2 patents and written numerous reports on development and analysis for a variety of applications. I began my consulting with criminal type analysis and reporting in 2000 and in 2005 opened 3rd Degree Investigations, Inc., where I currently have a mobile lab station for certified weight analysis and reporting. I have, over the last nine years, reviewed and consulted on hundreds of various types of analysis including, but not limited to DNA, Narcotics, Poisons, Toxic Metals, Crime Scene Analysis and Evidence Collection Procedures, and Pharmaceutical Products. In 2009, when I had successfully completed more than 5 years in the field of Forensics, I became eligible to be a member of the American College of Forensic Examiners Institute (ACFEI). The ACFEI is the world’s largest forensic science association, and it covers a broad range of forensic specialties. The association actively promotes raising awareness about forensic science and it supports its members as they work to advance their fields. ACFEI serves as the national center for this purpose and circulates information through its journal, The Forensic Examiner, lectures, seminars, conferences, workshops, continuing education courses and home study courses. Since being a member, I have completed the second level certification of Medical Investigator and have scheduled to attend the 2009 National Conference in October. I have testified in cases in numerous states, at the Military, Federal, State and Local levels on topics regarding DNA, Narcotics, Sexual Assault, and DUI. I have completed numerous case reviews; consulting and reporting on cases in numerous states on the topics of DNA, Narcotics, DUI, Sexual Assault, Gunshot wounds, Crime Scene Analysis and Evidence Collection Review and Reporting.

DNA Terms and Definitions:

DNA is the material that governs our inheritance of eye color, hair color, skin color, bone density, and many other human traits. DNA is a long string like object where one foot of this object can fit into a space of approximately 1/millionth of a cubic inch. Every part of our body is made up of tiny cells all containing strands of DNA. The only exception to this is red blood cells. DNA can be typed from blood due to the presence of white blood cells. Most living things including plants, animals, and bacteria rely on DNA. Each strand of DNA is made up of tiny building blocks that are sequenced differently for each living thing, allowing for differences and similarities to our relatives and others. Therefore DNA can be used as a tool to identify one person or one plant or one animal from another.

A locus is a specific location within the DNA strand. Each locus resides at a particular place within a chromosome. Chromosomes are simply bundles of DNA. For humans, there are 23 pairs of chromosomes, each with a consistent size and shape. Variations within the chromosome are called alleles. At each locus, a person has 2 alleles where 1 is inherited from each parent. It is possible for a person to have the only one allele detected whereby indicating that the person inherited the same allele from both parents. Each allele is a short fragment of DNA from a specific location on the human genome known as STR (short tandem repeats). STR’s are places in the human DNA where the genetic code repeats itself. Everyone has these repeating fragments where the number of repetitions and also the length of the segment vary among individuals.

Case Discussion:

Mr. Robert Reaves resides in Durham County with his sister and a male roommate named Mr. Steve Randolf. On January 29, 2008 an incident took place on the side of Interstate 540 near the Louisburg Rd. Exit involving Mrs. Latrese Curtis. Her deceased body is discovered on January 30, 2008, on the side of Interstate 540, the apparent victim of multiple stab wounds. Mr. Reaves became a suspect in this homicide investigation, as well his roommate Mr. Randolf. DNA samples were collected from the scene, including Mrs. Curtis’s vehicle as well her body. Her fingernails were reportedly broken and tissue fragments were recovered from underneath two of her fingernails (one on each hand). Evidence that was collected as well as reference samples from Reaves, Randolf, and Mr. Curtis (Latrese’s husband) were submitted to the SBI (State Bureau of Investigations) Laboratory for examination and analysis.

DNA evidence collected included swabs of the inside of Latrese Curtis’s white Nissan Sentra, fingernail clippings from Mrs. Curtis, tissue located on the fingernail clippings, various parts of Curtis’s body and the articles clothing that Mrs. Curtis was wearing at the time of her death. Reference DNA samples were obtained from the victim, her husband Darin Curtis and the possible suspects, Reaves and Randolf. Additionally, swabs were obtained from the fingernails of the hands of both Reaves and Randolf. In the STR/DNA analysis performed by the SBI Laboratory in Raleigh, NC, Reaves was excluded as a contributor from the swabs taken from Curtis’s Nissan Sentra; to include the driver seatbelt, passenger seatbelt, windshield wiper lever and driver seat adjustment bar. Mr. Reaves was also excluded as a contributor on the analysis of the fingernail clippings, tissue recovered from Curtis’s right and left hands, a vaginal swab of Curtis, the panties of Curtis, and a rectal swab of Curtis. A washcloth was obtained for examination from Reaves’s house where DNA analysis revealed a mixture that rendered “no opinion” for Reaves or Curtis as contributors and additional alleles present that could not be accounted for and did not match the reference samples provided. The DNA results of the back of Reaves’s hands resulted in NO MATCH to Curtis. The results of the steering wheel swab were consistent with a mixture with additional alleles present that could not be accounted for and did not match the reference samples provided. Reaves could not be excluded according to the lab report but was NOT listed as a MATCH.

Conclusion Discussion of Results:

A sample that contains DNA from 2 or more individuals is considered to be a mixture. Mixtures are the most common complication in the analysis of DNA evidence. A mixture is generally recognizable with more than 2 peaks resulting at any locus. By their very nature mixtures are difficult to interpret where the number of contributors is often unclear. Although the presence of three or more alleles at any locus is indicative of the presence of more than one contributor, it is often difficult to tell whether the sample originated from 2, 3, or even more individuals because the various contributors may share the same alleles. Refer to the Comparison of DNA Results table where at locus marker D8S1179 on the swab from the steering wheel of Curtis’s vehicle. Alleles are present at 13, 14, and 15. Looking at all of the reference standards these alleles may show that D. Curtis, L. Curtis, Randolf, and Reaves could also have been present. Both Mr. and Mrs. Curtis and Reaves all share the same values at marker D8S1179 and Randolf’s standard showed two alleles at 12 and 15 respectively. It is possible that Randolf could be considered a contributor at this marker based on the allele at 15 where the allele at 12 may have “dropped-out” from the analysis. There is no confirmation for “dropped” alleles and only subjective review for determining such. Another possibility is that there was another person present that had only one allele, 15, at this locus. There are other loci that show similar results among at least three of the standards reported. A study of one database of 649 individuals found over 5 million three-way combinations of individuals that would have shown the same four or fewer alleles across all 13 commonly tested STR loci.

Some laboratories attempt to identify or determine which alleles go with which contributor based on peak height. They assume the taller peaks go with the “major” or “primary” contributor, generally indicating larger quantities of DNA at the start of the analysis and the smaller peaks associated with a “secondary” or “minor” contributor. These inferences are sometimes problematic because a variety of factors other than the amount of DNA present can affect peak height. Sample degradation or break down can skew a relationship between peak heights. In mixed samples, it may be impossible to determine whether alleles of one or more contributors have become undetectable at some loci. Analysts have been known to be speculative on determining whether all the alleles are present or not, rendering alternative interpretations of the data. Furthermore, 2 or more samples that make up a mixture of DNA may have different rates of degradation, perhaps due to being deposited at different times or due to the differences in protection offered by different cell types. These possibilities render an interpretation of potentially degraded mixed samples particularly prone to subjective or unscientific interpretation. Additional complications with DNA analysis are spurious peaks, commonly referred to as “stutter” peaks (small peaks occurring immediately before and sometimes, although rare, after a real peak) and “noise” peaks (small background peaks that can occur along the baseline or bottom line of the run). A wide variety of factors including air bubbles and sample contamination can create small random flashes that occasionally may be large enough to be confused with an actual peak or even may mask an actual peak. Pull-up or bleed-through represents a failure of the analysis software to differentiate between different dye colors. Pull-up peaks are generally identifiable through careful analysis of the peak positions across the color spectrum. Duplicate runs of the same sample will often rule out many artifacts resulting in the run. Although many of these technical artifacts are clearly identifiable, standards for determining whether a peak is a true peak or a technical artifact are often rather subjective, leaving room for disagreement among experts.

Threshold issues are another area of concern in DNA analysis. Threshold values are the cut-off limit defining an actual peak value. A peak height (expressed in Relative Fluorescent Units or RFU’s) should exceed a certain value, determined by calibration of the analyzer, before it can be determined an allele. According to an interview with 2 different technical representatives at Applied Biosystems (manufacturer of genetic analyzers used by 85% of labs reporting results for DNA), each laboratory is responsible in determining their own peak thresholds. Applied Biosystems has been noted in recommending a peak threshold of 150 RFU and cautioning interpretations of peaks below this value. I interviewed a DNA Analyst with an independent laboratory holding similar certifications as the FBI Crime lab, uses a threshold of 150RFU. When the quantity of DNA being analyzed is very low (indicated by low peak heights) the genetic analyzer may fail to detect the entire profile of a particular contributor. These short peaks are referred to as “weak” alleles and are often difficult to determine whether they are low-level peaks or technical artifacts.

At locus D8S1179 Mrs. Curtis and Mr. Reaves share the same alleles of 13 and 14, as well does Mr. Curtis. The 13 and 14 peaks heights from the electropherogram (analysis run from the genetic analyzer) were 3166 and 2879 respectively on one of the duplicate submissions for analysis. The additional allele shown at this locus was 15 with a peak height of 371, noted as a “weak” allele. The 13 and 14 alleles at this locus, would by general standards, be considered the “major” contributor with the 15 allele belonging to a minor contributor. At locus TH01, again Mrs. Curtis and Mr. Reaves share the same alleles of 7 and 8, with peak heights at 3145 and 2517 respectively. There is an additional allele at this same locus of 9.3 with a peak height of 244, noted as a “weak” allele. It would be considered that the 7 and 8 allele combination would likely belong to the “major” contributor and the 9.3 belonging to a “minor” contributor. Along the entire electropherogram, Mrs. Curtis’s DNA appears to be prominent as a “major” contributor matching at least 9 of the 13 (a standard in DNA analysis interpretation recognized by the FBI Crime Lab) loci considered. The two loci points where Reaves and Curtis share allele combinations should be considered as just that, a shared combination. A DNA study of 649 individuals showed that over 5 million 3-way combinations would demonstrate up to four common alleles across all 13 commonly tested STR loci. In a population of 5 million people, 649 could potentially share up to 4 of the same alleles at any particular loci, which is why the standard for identity match is nationally recognized at a minimum of 9 matches of 13 loci observed. Relating this data to the population of Wake County, North Carolina, up to 112 people in the county could share the same alleles at up to 4 loci markers.

Referring to the table of DNA Comparison attached to this report, Mrs. Curtis reference sample predominately showed results with strong peak heights at all 13 loci observed. Present at loci D21S11 were 5 alleles 28, 36, 31, 31.2, and 32.2 with 28 and 36 having the dominate peak heights for a “major” contributor, leaving potentially the balance of noted weak alleles for interpretation as “minor” contributors. Finally, there is a weak peak at the AMEL loci indicating the presence of a male contributor from a “weak” allele peak.

Mr. Reaves shared alleles with Mrs. Curtis at 2 loci, where it is determined that Mrs. Curtis’s DNA is the “major” contributor. At 3 other loci there are weak alleles that may be considered as being similar to Mr. Reaves, however they are very weak and some are below 150RFU (the recommended threshold by Applied Biosystems, the manufacturer of the genetic analyzers used by 85% of labs reporting DNA results). Assuming again that the alleles that are shared at 2 loci belong to Curtis, the remaining 3 loci reviewed with weak alleles are not enough scientific data to determine presence or contact with the surface examined. A 3 loci match of 13 is not strong enough data to be considered as a contributor. Including the 2 loci that have shared alleles, there are only 5 loci that may be matching, which is still not strong enough to consider as a contributor. This is due to the very low peak thresholds where any peak below 150RFU should be interpreted with caution, such as some peak values at marker D21S11, where 2 of those peaks are below 150RFU. Since laboratories have the authority to determine their own threshold limits, another laboratory could have examined and analyzed the same data with an “eliminated” or “excluded” determination for Mr. Reaves. If we review the given data and remove all peaks below 150RFU, we would then only rely on 4 loci matches, that renders a result in the category referenced above, where 649 of 5 million or 112 of 866 thousand, can share up to 4 similar allele combinations at various loci.

DNA profiling cannot determine whether a person is the source of the DNA, but rather the likelihood of which two people have the same profile. The DNA analysis is only conclusive in determining a probability match of a randomly selected database. The FBI uses a database of all DNA profiles presented into CODIS (Combined DNA Index System). “The National DNA Index (NDIS) contains over 7,261,604 offender profiles and 277,215 forensic profiles as of July 2009.” North Carolina database for DNA profiling has approximately 161,925 offender profiles and 4022 forensic profiles.

Mr. Reaves was noted in the DNA analysis report completed by SBI Laboratory, as not being able to be excluded from item #12, identified as “swab of steering wheel”. The probability estimate was reported as being 1 in 147 thousand for the Black population. In representative terms, if in a group of 866 thousand people, Mr. Reaves would be 1 of 5 or 6 that could also be included. The profiles in the listed databases are representative of a proposed entire population. This is all done in a statistical manner based on how many occurrences there have been for various allele combinations present in the given profiles for given groups collected. After reviewing profiles for Caucasian, Black, Lumbee, and Hispanic on a random selection and obtaining similar data over those groups, a random probability calculation can then be determined for profiling DNA among the entire state population. There are approximately 866,000 people in Wake County, North Carolina and therefore if we divide 147,000 into 866,000 we would ascertain that any one in 5.89 people could have a similar profile, for at least up to four loci, as Mr. Reaves in Wake County. Therefore, there are approximately 1 in 5 chances that an innocent person could be linked to the DNA present on Mrs. Curtis’s steering wheel.

Disclaimer: While every effort has been made to ensure the accuracy of this publication, it is not intended to provide legal advice as individual situations will differ and should be discussed with an expert and/or lawyer.For specific technical or legal advice on the information provided and related topics, please contact the author.

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