ABSTRACT
AIM: Several amplification and detection formats for the analysis of short tandem repeat loci are readily available to the forensic laboratory. Careful consideration must be given to the throughput, sensitivity, concordance, data interpretation, facility requirements, and costs of operation. The Pennsylvania State Police DNA Laboratory sought to establish that of any of the amplification or detection formats generally used in the United States generates concordant results and that the use of several formats within one laboratory provides a solution to the interpretation of difficult evidentiary samples. METHODS: Validation work consisting of sensitivity, precision, mixture, and substrate studies was performed by use of each of three detection formats (ABI Prism(r)310 Genetic Analyzer, ABI Prism(r)377 DNA Sequencer, and the Hitachi FMBIO(r)II Fluorescent Scanner) and three amplification systems (GenePrint(r) PowerPlex 16, GenePrint(r) PowerPlex 1.1/2.1, and AmpflSTR ProfilerPlus/COfiler). The results generated in each of the formats were compared, along with the problems incurred. RESULTS: All allele calls were concordant, with the exception of primer region variants, and all detection systems were sensitive and reliable. Even with the use of multiple formats, a general protocol can be written with only one set of interpretation guidelines. CONCLUSION: National databases can be used with input data from any of these formats. The use of several detection formats allowed the forensic scientist to select a system, based on sample quality, quantity, and throughput requirements. Interpretation issues resulting from complex mixtures, degraded samples, rare microvariants, internal primer variants, unusual heterozygote ratios, above or below ladder alleles, and potential tri-alleles can be verified.
Subject(s)
DNA Fingerprinting/instrumentation , DNA Fingerprinting/standards , Forensic Medicine/instrumentation , Forensic Medicine/standards , Nucleic Acid Amplification Techniques/instrumentation , Nucleic Acid Amplification Techniques/standards , Criminology/methods , DNA Fingerprinting/methods , Female , Fluorescent Dyes , Forensic Medicine/methods , Humans , Male , Pennsylvania , Polymerase Chain Reaction/instrumentation , Polymerase Chain Reaction/methods , Sensitivity and Specificity , Tandem Repeat SequencesABSTRACT
Interpretation guidelines for short tandem repeat casework analysis are difficult to construct. As soon as a set of guidelines are developed, a new case evolves that does not fit the painstakingly written document. The casework analysts gather and amend the guidelines again, and again. This article seeks to demonstrate that general guidelines can be set and written such that it can be used for any detection format. Guidelines published by the Scientific Working Group for DNA Analysis Methods, a working group of DNA forensic experts in the United States, are used to set the format for the written protocol on interpretations. The rule "the interpretation of results in casework is a matter of professional judgment and expertise. Not every situation can or should be covered by a preset rule" is stressed. Development of minimum and maximum threshold values, heterozygote ratios, stochastic limits, and determination of major and minor components based on validation studies is discussed. The paper travels through setting criteria to evaluate internal lane standards and amplification controls. It continues with establishing ranges for interpretation and defining true alleles versus anomalies. Examples of a variety of profiles are given and the potential interpretation, using signal intensities and genetics. In addition, report writing strategies and wording routinely used by the Pennsylvania State Police DNA Laboratory System are given.
Subject(s)
DNA Fingerprinting/standards , Forensic Medicine/standards , Guidelines as Topic , Tandem Repeat Sequences , Alleles , Female , Humans , Male , Nucleic Acid Amplification Techniques , Sensitivity and Specificity , United StatesABSTRACT
Statistical analysis was performed on a subset of the Pennsylvania State Police Caucasian, African American and Hispanic database for the purpose of determining Hardy-Weinberg equilibrium and associations across the RFLP loci D1S7, D2S44, D4S139, D5S110, D10S28 and D17S79 and the PCR-based loci HLA-DQA1, LDLR, GYPA, HBGG, D7S8 and Gc. Overall, the statistical results are consistent with a population in equilibrium both within and between loci. The assumption for independence is valid.
