A bioinformatic tool was developed to assist with the victim identification initiative that followed the Swissair Flight 111 disaster. Making use of short tandem repeat (STR) DNA typing data generated with AmpFlSTR R Profiler Plus TM (PP) and AmpFlSTR R COfiler TM (CO) kits, the software systematically compared each available STR genotype with every other genotype. The matching algorithm was based on the search for: (i) direct matches to genotypes derived from personal effects; and (ii) potential kinship associations between victims and next-of-kin, as measured by allele sharing at individual loci. The software greatly assisted parentage analysis by enabling kinship evaluation in situations where complete parentage trios were unavailable and, in some situations, with distantly related relatives. Exclusion of fortuitous kinship associations (FKA) was made possible through the recovery at the disaster site of at least one remains for every sought-after victim, and was incorporated into the kinship software. The data from the 13 combined STR loci produced 6 and 23 times fewer FKAs when compared with PP alone and AmpFlSTR R Profiler TM (PR) alone, respectively. Identification leads or confirmations of identification were obtained for 218 victims for which DNA reference samples (personal effects and kin) had been submitted. Confirmation of an inferred kinship association was sought through frequency and likelihood calculations, as well as corroborative data from other identification modalities. The use of a simple, yet powerful, automated genotype comparison approach and the use of megaplexes with high power of discrimination (PD) values extended considerably the identification capabilities in the case of the Swissair disaster. The DNA typing identification modality proved to be a valuable component of the large arsenal of identification tools deployed in the aftermath of this disaster.

To assist the interpretation of STR DNA typing results from forensic casework samples containing mixtures, the range of heterozygous allele peak height and peak area ratios (HR) and stutter percentages (stutter %) for the loci comprised in the AmpF STR R Profiler Plus TM (PP) kit were assessed on 468 database and 275 casework single source samples. Stutter % medians were similar for database and casework samples, ranging from 2% to 7%. The upper limit of the stutter value range was 16%, calculated as median +3 SD, although lower locus-specific values could be used. HR medians were 93 ± 6.5% for database samples, 88 ± 12% for casework samples. For casework samples, the maximum signal imbalance noted was 52%, calculated as median −3 SD. No significant difference was observed between peak height and peak area calculated values. This study shows the importance of selecting the proper reference database for the establishment of HR threshold values.

Two major goals in the development of new DNA typing technology for forensic use have always been: (1) to achieve the highest power of discrimination possible using a valid and reliable method, and (2) to preserve limited evidentiary samples by utilizing the smallest portion possible for analysis. The discovery of the STR markers (1-3) as well as the development of fluorescence detection instruments (4-9) provided forensic scientists with sophisticated means of achieving their original goals. The development of highly discriminatory megaplex STR systems used in a fluorescence-based detection mode significantly increased the capability for analyzing a larger number of samples that have a limited amount of biological material (10-16). This combined approach offered great sensitivity and accuracy as well as facilitated the interpretation of DNA mixtures by providing data, such as peak height and peak area, that can be integrated into formulas and subsequently analyzed to assist in profile interpretation (17-20). Many validation studies have been published that indicate that STR systems, in multiplex formats, provide excellent typing results for samples subjected to a variety of environmental and experimental conditions (15,21-27). Despite this valuable information, newly developed STR multiplex systems must be subjected, by the practicing forensic community, to a thorough examination in order to define their limitations and establish their robustness and reliability under specific circumstances and/or experimental conditions. Only from such studies can scientifically sound interpretation guidelines be derived and a universal consensus for data sharing among different forensic laboratories be obtained. This report presents some of the studies that were conducted during the validation of the AmpFᐍSTR ® Profiler Plus™ STR amplification system using reduced PCR volume conditions (i.e., 25 L). Although the manufacturers of the AmpFᐍSTR ® commercial kits recommend the use of 50 L as the PCR volume (15,28), previous experience gained using STR multiplexes under reduced PCR volume conditions (21,29) and our involvement in the FBI STR Standardization Project suggested the evaluation of the amplification conditions described in this report. Six different categories of Profiler Plus™ profiles were defined following the processing of 275 biological evidence in 25-L PCR volume. Examples for each category are presented along with data collected and used to develop our interpretation guidelines. As a means to improve the odds of obtaining balanced and complete profiles for casework samples showing partial profiles or profiles with a slope, several amplification conditions are presented. In addition, a series of simulated mixtures representing ten different mixture scenarios was prepared to establish the limit of detection of a minor profile using the Profiler Plus™ multiplex system. Two different amounts of total DNA (1 and 2 ng) and different ratios ranging from 1:20 to 20:1 were selected to cover a wide range of potential casewo...

HumD21S11 and HumFGA (multiplex 1B) have been evaluatedWe reported earlier (24,25) the development of a fluorescencefor use in forensic identification using the Applied Biosystems based multiplex DNA profiling system comprising two highly Model 373A and Prism 377 DNA Sequencers, respectively. Experiments were aimed at defining the limit of target DNA polymorphic STR loci, HumD21S11 (26) and HumFGA (27), and required for reliable profiling, the level of degradation that would the amelogenin gender determination system (28,29). In the present still permit amplification of the short tandem repeat (STR) loci report, we document the extensive validation studies performed examined, and the robustness of each locus in the multiplexes after and casework experience gained using this triplex system on the samples were exposed to environmental insults. In addition, the Applied Biosystems Model 373A DNA Sequencer. Validation specificity of the multiplexes was demonstrated using nonhuman DNAs. Forensically relevant samples such as cigarette butts, chewexperiments were also conducted using a modified triplex comprising gum, fingernails and envelope flaps were processed using both ing HumD3S1358 (30), HumD21S11 and HumFGA on the secondan organic extraction procedure and a QIAamp protocol. DNAs and generation ABI Prism 377 DNA Sequencer. Initial experiments resultant multiplex STR profiles were compared. The validation of were designed to determine the optimal amount of target DNA for the triplex STR systems was extended to include over 140 nonproreliable amplification results on both analytical instruments. As a bative casework specimens and was followed with a close monitoring of initial casework (over 300 exhibits). Our results document complement to previous reports documenting the suitability (quanthe robustness of these multiplex STR profiling systems which, tity and quality) of DNA extracted from specimens under extreme when combined with other multiplex systems, could provide a environmental conditions for RFLP profiling (31-34) or power of discrimination of approximately 0.9999. 16,[35][36][37][38][39][40], this paper further examined adverse conditions relevant to fluorescence-based amplifica-