page_title: Forensic biology
section_title: DNA analysis
hierarchical_section_title: Forensic biology / Disciplines / DNA analysis
caption_attribution_description: English: An electropherogram of a three person mixture in a 5:2:1 ratio using the Applied Biosystem's Identifiler STR kit
context_page_description: Forensic biology is the application of biology to associate a person, whether suspect or victim, to a location, an item, another person. It can be utilized to further investigations for both criminal and civil cases. Two of the most important factors to be constantly considered throughout the collection, processing, and analysis of evidence, are the maintenance of chain of custody as well as contamination prevention, especially considering the nature of the majority of biological evidence. Forensic biology is incorporated into and is a significant aspect of numerous forensic disciplines, some of which include forensic anthropology, forensic entomology, forensic odontology, forensic pathology, forensic toxicology. When the phrase "forensic biology" is utilized, it is often regarded as synonymous with DNA analysis of biological evidence.
context_section_description: DNA, or deoxyribonucleic acid, is one of the most popular pieces of evidence to recover at a crime scene. More often than not, evidence containing DNA is regarded to as biological evidence. With all of the substantial advances that have been made regarding DNA, biological evidence is recognized to be the golden standard in forensic science.
  At the scene, biological evidence must be initially visibly recognized. Sometimes this is not always possible and the aid of an alternative light source, or ALS, is required. Once identified as a potential source, presumptive tests are conducted to establish the possibility of the specified biological presence (semen, saliva, blood, urine, etc.). If positive, samples are collected and submitted for analysis in the laboratory, where confirmatory tests and further tests are performed.
  DNA analysis has numerous applications such as paternity testing, identifications of unknown human remains, cold case breakthroughs, as well as connecting suspects and/or victims to a piece(s) of evidence, a scene, or to another person (victim or suspect, respectively). Nuclear DNA evidence can be recovered from blood, semen, saliva, epithelial cells and hair (if the root is still intact). Furthermore, Mitochondrial DNA can be recovered from the shaft of hair, bone and the roots of teeth. For most forensic DNA samples, STR analysis of autosomal short tandem repeats is performed in an attempt to individualize the sample to one person with a high degree of statistical confidence.
  Laboratory analysis of DNA evidence involves the sample DNA being extracted, quantitated, amplified, and visualized. There are several methods of DNA extraction possible including organic (phenol-chloroform) extraction, Chelex extraction, and differential extraction. Quantitation is commonly conducted using a form of the polymerase chain reaction, known as real-time PCR, quantitative PCR, or qPCR. qPCR is the preferred method of DNA quantitation for forensic cases because it is very precise, human-specific, qualitative, and quantitative. This technique analyses changes in fluorescence signals of amplified DNA fragments between each PCR cycle without needing to pause the reaction or open the temperature-sensitive PCR tubes. In addition to the components necessary for standard a PCR reaction (i.e. template DNA, carefully designed forward and reverse primers, DNA polymerase [usually Taq], dNTPs, and a buffer solution containing Mg2+), qPCR reactions involve fluorescent dye-labelled probes that complement and anneal to the DNA sequence of interest that lies between the two primers. A “reporter” (R) dye is attached at the 5’ end of the fluorescent probe while a “quencher” (Q) dye is attached at the 3’ end. Before the DNA strands are extended by the polymerase, the reporter and quencher are close enough in space that no fluorescence is detected by the instrument (the quencher completely absorbs/masks the fluorescence of the reporter). As the polymerase begins to extend the strand, the 5' end of the probe is degraded by the polymerase due to its exonuclease activity. The reporter dye is released from the 5’ end and is no longer quenched, thus enabling detection of fluorescence. A graph is constructed for the sample DNA comparing the presence of fluorescence (y-axis) to cycle number (x-axis) of the qPCR process. This is then compared to a standard curve of the cycle fluorescence threshold (y-axis) versus the log of known DNA concentrations (x-axis). By comparing the sample data to the standard curve, one may extrapolate the DNA concentration in the sample, which is essential to move forward with PCR amplification and capillary electrophoresis to obtain a DNA profile. DNA profiles are produced in the form of an electropherogram. The obtained profile can be compared to known samples such as those in CODIS in order to identify a possible suspect. Based on known frequencies of the genotype found in the DNA profile, the DNA analyst may place a statistical measure of confidence on DNA match.
caption: STR electropherogram of a three-person mixture
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