Análise genética de impressões digitais - Amostras Low Copy Number

Abstract

The possibility to perform low copy number DNA typing, when just a few cells are available, as changed the way how crime scene investigations is faced. Nowadays it is possible to successfully type some evidence that couldn t be considered until now. Fingerprints are a good example of those. Since that just a few cells are present in this evidence (enabling recovery of low quantities of DNA, fewer than 100pg) just very sensitive systems can detect nuclear DNA. The most used method is definitely increasing the number of PCR cycles. However, increased occurrence of stutters and artifacts that reduced the quality of the DNA profile is normally observed. The present work aimed to compare the application of autosomic STR, Y-STR and miniSTR markers, based on the concept of increased number of PCR cycles as a strategy to achieve more sensitivity. Some other methods, such as whole genome amplification and nested-PCR, were also evaluated as an alternative way to reach the desired sensitivity. Another goal was to determine the influence of several reagents for developing latent fingerprints (cyanoacrylate fuming, magnetic powder and white powder) in DNA typing. The results shows that increasing the number of PCR cycles still is the best way to attain the required sensitivity. Moreover we could realize that DNA was partially degraded, once there were observed significant differences between loci larger and smaller than 200bp. Among all markers miniSTR showed to perform the best results in terms of detected alleles percentage. On the other hand, Y-STR seemed to be highly affected in the presence of degraded DNA and PCR inhibitors, which makes them less robust for these analyses. LCN profiles are significantly affected by artifacts, like allele dropout and heterozygous imbalance, derived from stochastic fluctuation. Reporting consensus profiles reduces artifact inherent errors. Finally, cyanoacrylate proved to have a minimum negative effect on DNA profiling, while white powder was the worst reagent.