Browsing by Subject "Forensic Genetics, instrumentation"
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Item A Bone and Buccal Sensitivity Study Comparison and Stability Study using the PowerPlex[R] Fusion 6C System(2018-05) McDaniel, Ethan L.; Warren, Joseph E.; Planz, John V.; Krishnamoorthy, Raghu R.; Gaydosh-Combs, LauraA validation study, a bone sensitivity study and a stability study were performed using the PowerPlex[R] Fusion 6C System. These studies were performed on a 7500 Real-Time PCR System, 9700 GeneAmp Thermocycler and 3500xL Genetic Analyzer. Buccal DNA was used to develop a method to analyze the DNA profiles gathered during the bone sensitivity study and stability study. DNA profiles for specific concentrations of DNA in solution were obtained during the bone sensitivity study. The stability study showed profiles exhibiting the effects of metal PCR inhibitors being introduced to the DNA extract solutions. Full profiles were obtained for calcium concentrations less than 7.35 mM, while the instrument was fully inhibited for copper concentrations between 0mM and 7.35 mM. Based on the limited data, the PowerPlex Fusion 6C System cannot tolerate copper when present in DNA solutions; whereas, calcium may be tolerated as an inhibitor up to 7.35mM.Item A Validation Study of the Applied Biosystems[TM] GlobalFiler[TM] PCR Amplification Kit on the Applied Biosystems[TM] 3500xL Genetic Analyzer(2018-05) Cuthbert, Clare C.; Planz, John; Allen, Michael; Roane, Brandy; Warren, JosephThe main focus of this internship was the validation of the Applied Biosystems[TM] GlobalFiler[TM] PCR Amplification Kit on the Applied Biosystems[TM] 3500xL Genetic Analyzer. Internal validations are essential in accredited forensic laboratories, as it is required by Standard 8 of the FBI Quality Assurance Standards (2011) that new procedures, instruments and kits, have to be validated prior to their implementation into casework. According to the SWGDAM guidelines, the validation study has to include several studies; known and non-probative evidence samples or mock evidence samples, sensitivity and stochastic studies, precision and accuracy studies, a concordance study, mixture studies, a degradation study and a contamination assessment. At the Harris County Institute of Forensic Sciences (HCIFS), the validation of the Applied BiosystemsTM 3500xL Genetic Analyzer was conducted in the Genetics Laboratory. The 3500xL Genetic Analyzer has numerous enhanced capabilities over the older instrument that HCIFS currently uses for casework; the Applied BiosystemsTM 3130xL Genetic Analyzer. The 3500xL can analyze six dyes, has RFID, has 24 capillaries, and a solid-state long-life laser. The samples were extracted and purified for use on the 3500xL via QIAgen or the Organic method. The Quant Trio Kit, and the GlobalFilerTM PCR Amplification Kit were also used in the process. After injecting the samples on the 3500xL, they were analyzed appropriately for each study. 28 cycles for amplification and a 24 second injection time were used. An analytical threshold of 50 RFU for blue, 65 RFU for yellow, 45 RFU for green, 55 RFU for red and 60 RFU for purple were used, with a stochastic threshold of 400 RFUs. An optimal DNA amount of 0.5ng was also used. The results of each study showed that the 3500xL to give reliable and reproducible results, and the instrument performed as expected.Item ParaDNA: A Novel Instrument for Presumptive DNA Analysis(2018-05) Moussa, Ola; Warren, Joseph E.; Allen, Michael S.; Staub, Rick W.; Millar, J. CameronCurrent methods for human identification are time consuming and can take weeks to complete, leading to a backlog of evidence needing to be processed and a slowdown in investigations. The ParaDNA[R] Instrument is designed to address this issue using the Screening and Intelligence Systems which can detect the relative amount of DNA in an evidence item, and analyze 5 short tandem repeats (STRs) and Amelogenin, respectively [3]. The Instrument uses HyBeacons[R] which target specific STRs to identify the presence of DNA, and detect STR alleles. This is a validation of the ParaDNA[R] Screening and Intelligence Systems using saliva and blood samples to assess the sensitivity and reliability of the instrument. The data collected using the ParaDNA[R] Instrument show that it can reliably identify the relative amount of DNA in a sample, and display useful STR profiles that are 99% concordant with Qiagen Investigator 24Plex QS STR Kits.