The use of higher PCR cycle numbers (e.g., 36 or 42) and the already amplified copy number of mtDNA per cell necessitate great care to avoid contamination. The DNA templates under investigation are often damaged so they may not be as readily amplified as even low amounts of high quality DNA from laboratory
Process for evaluation of mtDNA samples. The evidence or question (Q) sample may come from a crime scene or a mass disaster. The reference or known (K) sample may be a maternal relative or the suspect in a criminal investigation. In a criminal investigation, the victim may also be tested and compared to the Q and K results.
Performed separately and preferably after evidence is completed
personnel or reference samples. Reference samples from the victim, the suspect, and maternal relatives are typically available as blood stains or buccal swabs and generally contain large amounts of high quality DNA.
Practices to reduce or minimize contamination often employed by forensic laboratories performing mtDNA testing include use of protective clothing such as disposable lab coats, frequent cleaning procedures with bleach and UV irradiation of hoods and lab bench surfaces, processing the question samples prior to the known samples, multiple glove changes during sample handling, using dedicated equipment for the mtDNA testing, and physically separating the pre- and post-amp spaces. During an analytical procedure only one item of evidence from a case is opened at a time (Isenberg and Moore 1999). Some laboratories even control movement of laboratory personnel between spaces. For example, a technician may not be permitted on the same day to return to a pre-amplification area after having entered a post-amp area. Vigilance on the part of all laboratory personnel is important to keep a forensic mtDNA laboratory clean. Reagent blanks and negative controls are also run to monitor levels of exogenous DNA in reagents, laboratory environment, or instruments.
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