Tissue-Specific DNA methylation patterns in forensic samples detected by Pyrosequencing®

In certain circumstances the outcome of a trial may hinge on the ability of a forensic laboratory to determine the identity of biological stains present at crime scenes. An example of such a situation would be the detection of blood, saliva, vaginal fluid, or other body fluid in a specific location whereby its presence would reinforce the victim’s or suspect’s version of the events that happened during the commission of a crime. However, current serological methods used for identifying body fluids may lack the sensitivity and specificity to identify these fluids, particularly for trace levels. New procedures using proteomic methods and RNA-based gene expression show promise in addressing this issue; however, concerns about stability and relative levels of gene expression remain. An alternative approach is to utilize patterns of epigenetic DNA methylation. DNA methylation is an epigenetic mechanism that regulates the specificity of genes being expressed or silenced in cells. Regions in the human genome referred to as tissue- specific differentially methylated regions account for unique patterns of DNA methylation that are specific for each cell type. This chapter addresses the application of bisulfite-modified PCR combined with Pyrosequencing ® to detect tissue-specific DNA methylation patterns and perform trace serological analysis. The quantitative nature and precision available with Pyrosequencing presents major advantages in these studies as it permits detection of and contrast between cells with differential levels of methylation. The procedure can be applied to a variety of biological fluids which may be present at crime scenes.