FORENSIC SCIENCES: CRIMINALISTICS - Bio Medical Forensics

A162 Improved Genotyping Performance of
Complex Multiplexes on DNA Mixtures
Chien-Wei Chang, PhD*, Life Technologies, 850 Lincoln Centre Drive,
MS 404-1, Foster City, CA 94404; Robert Lagace, BS, Applied
Biosystems, 850 Lincoln Centre Drive, Mail Stop # 404-3, Foster City, CA
94404; Dennis Wang, PhD, and Julio J. Mulero, PhD, Applied
Biosystems, 850 Lincoln Centre Drive, MS 404-1, Foster City, CA 94404;
Lisa M. Calandro, MPH, Applied Biosystems, 850 Lincoln Centre Drive,
Foster City, CA 94404; and Lori Hennessy, PhD, Life Technologies, 850
Lincoln Centre Drive, MS404-1, Foster City, CA 94404
After attending this presentation, attendees will understand the
performance of complex STR multiplexes in DNA mixtures and the
parameters influencing mixture interpretation.
This presentation will impact the forensic science community by
providing information on factors that affect the performance of different
STR multiplex systems in compromised samples and solutions to
improve the recovery of information from complex multiplexes.
Evidence submitted for DNA analysis can be recovered from a
variety of biological samples including blood, saliva, or semen stains on
different substrates, body surface swabs, hair, bones, and finger nail
scrapings. Forensic analysts seek technologies to maximize the quality of
results obtained and increase the success rate of obtaining a DNA profile
from these types of samples. Multiplex short tandem repeat (STR) assays
simplify genotyping processes and preserve sample by eliminating the
need for multiple amplifications and/or electrophoretic injections.
Observations have previously been presented that there is varying
performance with different STR multiplexes. Therefore, a study was
undertaken to investigate the effect of multiplex complexity on
genotyping performance in mixtures by comparing STR assays of
different complexity (e.g., 10-16 markers/reaction) using simulated
inhibited samples at various mixture ratios (1:0, 1:1, 1:5, and 1:7).
Amplifications were performed with a total DNA input of 1.5 ng in either
1X TE, 15µM hematin, or 6ng/µl humic acid concentrations, chosen to
provide a moderate level of inhibition and generate a pattern
characteristic of inhibited profiles. Allelic drop-out of the minor alleles
in prepared inhibited samples was observed in all of the assays at each
mixture ratio; but when compared to the 10-plex assay, the 16-plex assay
detected an equivalent or greater number of alleles from the minor
contributors in all the mixture samples. Comparison of the intra-locus
balance values of the major and minor contributors were similar for all
assays. Some loci exhibited greater sensitivity to PCR inhibition in a
larger multiplex so modifications were made to the PCR buffer and
thermal cycling parameters in an effort to improve performance of these
loci. The improvements in PCR buffer and cycling conditions enabled
recovery of all alleles in the inhibited mixtures amplified using the 16plex
assay and eliminated the ski slope effect seen with the other kits
tested. This study illustrates how large multiplexes can be optimized
successfully for use on challenging casework samples.
Multiplex, STR, Mixture
* Presenting Author
A163 Increasing STR Analysis Success Rate and
Potential Discrimination With
Improved Multiplexes
Martin Ensenberger, PhD, Patricia M. Fulmer, PhD, Benjamin Krenke,
MS, Robert McLaren, PhD, Dawn R. Rabbach, AS, Cynthia J. Sprecher,
BS, Jonelle Thompson, MS, Douglas R. Storts, PhD, and Erin McCombs,
BS*, Promega, 2800 Woods Hollow Road, Madison, WI 53711
After attending this presentation, attendees will understand potential
strategies for handling challenged samples and increasing discrimination
for difficult relationship testing using an increased number of loci.
This presentation will impact the forensic science community by
demonstrating capabilities of new typing strategies.
Multiplex short tandem repeat (STR) analysis remains the primary
technique for human identification. At the beginning of this decade, focus
of STR multiplex design was on increasing the number of concurrently
analyzed markers, largely to meet the demand of having the FBI CODIS
core 13 loci in a single assay. Forensic analysts require STR systems that
are compatible with ever more challenging samples, prompting the need
for greater performance from assays. Additionally, the complexity of
relationship testing for immigration and familial searches has prompted
the need for increased marker availability. This combination of greater
performance and increased marker availability has driven the design of
the most recent generation of STR multiplexes.
Forensic samples routinely include impurities known to inhibit PCR
and reduce genotyping success rates. Additionally, high heat and other
environmental impacts can reduce the integrity of the DNA. Improved
buffer systems and incorporation of shorter amplicon primer design (mini
STR) have significantly increased the tolerance to common inhibitors and
yield from degraded samples. Additionally, increased sensitivity can
improve the likelihood of obtaining interpretable data from low
concentration samples and challenging mixtures.
The recommendation to extend the current European Standard Set
(ESS) for STR systems has prompted inclusion of several new markers in
the latest STR multiplex designs. Coamplification of two multiplexes can
provide full CODIS and ESS panels, plus Amelogenin, SE33, Penta E,
and Penta D, for a total of 24 markers. Compared to the CODIS core
panel, the additional markers add significantly to the power of
discrimination that can be applied to statistically-challenging cases.
Comparison data of these systems will be presented with inhibitors
and challenging samples along with developmental validation data. We
will also present strategies for use of these newer STR systems in the
forensic laboratory.
STR Analysis, Mini STR, Inhibition
A164 Human Genetic Identification in Cases of
Uncertain Kinship
Andrea Pinzon, MSc*, Instituto de Medicina Legal, Calle 4B Nº 36-01,
CALI, COLOMBIA
After attending this presentation, attendees will understand how in
forensic genetics cases, human identification is an important tool to relate
victims to their closest blood relatives. This requires highly polymorphic
molecular markers in order to find probable kinship between the victim
and his/her family members with a high level of certainty. The purpose
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