Collection of Biological Evidence from Pipe Bomb
The analysis of genetic markers containing short tandem repeats (STRs) has become the mainstay of the forensic DNA community to solve crime. DNA is extracted from biological evidence and multiple STR markers (or loci) are amplified using the polymerase chain reaction (PCR). The PCR process allows for the routine analysis of DNA quantities as low as 200 picograms of DNA (200 X 10-12 grams of DNA). However, in some cases the amount of DNA that is recovered from the evidence is limited; especially in cases involving touched items of evidence.
Capillary Electrophoresis of STR Fragments with Kaylie Slaughter (Master’s 2012)
The use of alternate techniques to increase the sensitivity of STR analysis is often employed in these cases by forensic laboratories across the country and around the world. These techniques can be as simple as increasing the amount of product loaded onto instruments, or can be as aggressive as increasing the number of PCR cycles (and therefore the sensitivity) in the amplification process. As a result, STR typing can be routinely performed on samples with as little as 10-20 picograms of DNA; 1-4 cells worth of DNA. A cautionary note is required here, as proper validation studies must be conducted prior to performing low-template testing to ensure the reliable reporting of data.
Our laboratory is studying the effects of increasing the sensitivity of STR typing procedures when using the STR kits and instruments available today; for example the PowerPlex 16 kits from Promega and the Identifiler/Plus kits from Applied Biosystems run on the Applied Biosystems 3130xl capillary electrophoresis instrument. We have altered the instrument conditions (amount of PCR product loaded, and the time and voltage of injection of product onto the instrument) and have increased the PCR cycling numbers (from 28 cycles to 31 or 34 cycles), and are studying the effects of changing these conditions on the interpretation of STR results. What is clear is that revised interpretation parameters are required for the analysis of STR data when low-template conditions are employed. Repetitive testing on the same sample, altering the thresholds for acceptance of data, and conservative statistical approaches for measuring the weight of the results are all important considerations when performing low-template STR analysis.