FORENSICS: HOW DOES DNA PROFILING WORK?

Although all humans are 99.9% genetically identical, there are some varying parts of the DNA used to distinguish between individuals. A DNA sample is analyzed to procure the specific pattern, called a profile, of a person's genetics in a process called DNA profiling. It is commonly used in criminal investigations to compare suspects’ DNA with that of samples obtained from the crime scene; a match between samples taken from the crime scene and the suspect could mean the innocence or guilt of the person. It is truly fascinating to consider that it has only been a mere 33 years since the technique was first used to convict a criminal, Colin Pitchfork, sentenced to life in prison in January 1988, guilty of rape and homicide of two teenage girls. While this process was revolutionary in its time of creation, it is now a global technique depicted in most crime-related tv shows. In a survey done by INTERPOL (The International Criminal Police Organization), it was found that 89 of the 110 countries that responded use DNA profiling in their criminal investigations. Since its first use, it has led to the conviction of millions of criminals and a lasting change in forensics. So how does this groundbreaking and surprisingly recent technique work exactly?

DNA profiling is used to compare several people's genetic information based on the number of repeats of specific sequences in our DNA. It can be briefly explained in a few paragraphs, covering everything from DNA Extraction to finding a match. However, before diving into the specifics, it is essential to review the basic DNA structure to understand the process.

Due to its ability to form complex molecules through covalent bonds, carbon is considered the foundation of life. Any compound within a living being and containing carbon is called an organic compound. DNA is composed of nucleotides, an organic compound containing a phosphate group, a base, and a 5-carbon sugar called deoxyribose.

Figure 1: Nucleotide, Wikipedia

These nucleotides are linked together by covalent bonds between the phosphate group of one nucleotide and the pentose sugar of the other nucleotide to form a long vertical strand. Two strands are also connected, as nitrogenous bases are attracted to each other by hydrogen bonds, linking together the two strands of DNA to form an antiparallel and double helix structure. This means that the two strands run parallel in opposite directions, winding around each other. Only certain types of bases attach to each other; this is called complementary base pairing. Adenine is complementary to Thymine, and Cytosine is complementary to Guanine due to their energetic favorability.

Figure 2: Organisation of DNA, BioNinja

Complementary pairings allow the DNA to replicate semi-conservatively, where an enzyme called Helicase is used to unwind the double helix and break the hydrogen bonds between strands. Using complementary base pairing, free nucleotides are linked to each of the now separate strands to form two whole new strands, double the amount of genetically identical DNA. The term ‘semi-conservative’ refers to how half the new DNA is formed from the original DNA.

DNA Profiling uses the same concept of semi-conservative replication, applying it to an artificial setting where all the conditions required for replication are met using equipment. In order to produce accurate results, scientists must first ensure that DNA samples are collected per an established set of rules, where specific steps are taken to minimize the probability of contamination of a sample, including the use of personal protective equipment and specialised collection kits. Great care is taken during transport, and storage is also at room temperature and away from any moisture to preserve the sample’s structural integrity.

To construct a DNA profile, scientists only require a small amount of DNA; a few cells suffice. The process is based on a non-coding region of DNA called Short Tandem Repeats (STRs), where the same order of nucleotides is repeated in a strand. This is crucial to DNA profiling as the number of repeats vary drastically from one person to another, allowing scientists to distinguish between individuals. In order to produce enough DNA for analysis, scientists use Polymerase Chain Reaction (PCR) to make millions of artificial copies of the DNA region containing STRs.

PCR can be broken down into three crucial steps:

1. Denaturing. The DNA sample is heated so that it denatures, breaking the hydrogen bonds between complementary bases and separating them into two single strands.

2. Annealing. Temperature is then lowered to one optimal for primers, a short strand of nucleotides, to bind to each strand to act as a beginning for the construction of an entirely new strand.

3. Extension. A heat-resistant enzyme called ‘Taq Polymerase’ is used to link the free nucleotides to the primers, extending it to make a complete double-strand structure, hence the process of extension. This is done using the pre-existing DNA strand as a template to provide complementary bases, resulting in double the amount of DNA than in the original sample.

Several cycles of heating and cooling allow the process to be repeated enough to produce millions of copies and enough to be analyzed. During the process, a fluorescent dye is used to tag DNA fragments so that the pattern of repeats is more easily distinguishable to the eye. The next step is to arrange the fragments of DNA obtained by size to compare two different DNA samples.

Figure 3: Polymerase Chain Reaction by Jodi So, CK-12 Foundation

The DNA sample is put through a capillary electrophoresis machine, which separates DNA fragments according to size. DNA is charged, so when an electric field is applied to it, the molecules will move. Usually, this happens at an equal rate for all molecules when DNA is submerged in liquid; however, the DNA sample is submerged in agarose gel, which allows the fragments to move at different speeds due to its consistency. Smaller fragments move faster than larger fragments, resulting in the DNA being rearranged by size.

Figure 4: Gel Electrophoresis by Khan Academy

Once the DNA undergoes gel electrophoresis, the fluorescent dyes added during PCR allow the fragments to be distinguishable, appearing as bands. The whole process of DNA profiling is carried out on both samples found on the crime scene and from the main suspects. The profiles of each are then compared, and those with bands that appear to be very similar suggest that both samples originate from the same individual.

Figure 5: Multiplex polymerase chain reaction (PCR) results for Monilinia spp. isolates from plum (Note: The numbers at the top represent the samples, while the ones on the left and right are a measure of the size of the fragments)

A DNA match means the person tested was at the crime scene for any reason before or after the crime. DNA can be found virtually anywhere on a crime scene. Traces of saliva can be found on cups, Tupperware, or cigarettes. If a crime involves a close struggle between victim and perpetrator, the DNA of the perpetrator may be found under the victim’s nails. For someone to be convicted, there must be a reason to believe that the collected sample is from the perpetrator and not anyone else. Nowadays, convictions require more than just DNA evidence; however, it may go a long way in convincing the jury at a court because it is a very well-established scientific technique.

While DNA profiling requires meticulous care in collection and transport, it is genuinely a fantastic technique when done right. It has revolutionized forensics and criminal investigation and became a technique heavily relied upon by many governments and institutions. It has also been used by institutions such as the Innocence Project to exonerate many falsely convicted people in hopes of reforming the system. It has many applications beyond forensics, including matching unidentified bodies, perhaps disfigured, to an identity and choosing organ donors that pose the slightest chance of rejection from the patient's body. It is also used in paternity tests and reuniting missing family members in wartime.