## DNA holds the answer

The main role in forensic genetics is played by the DNA molecule, the structure of which is described in more detail in Chapter 3 - Meet DNA, the bearer of genetic information. At this point, we must remember that DNA has to be packaged very precisely to fit into the cell nucleus. Several important proteins (e.g. histones) participate in the process of wrapping DNA and compaction, and DNA together with proteins forms a structure known as a chromosome. A human somatic cell has 46 chromosomes (23 pairs). Those marked with numbers are called autosomes and besides them, we have 2 sex chromosomes (gonosomes), X and Y. Females have two X chromosomes, and males have an X and a Y. We have two copies of each chromosome, one inherited from our mother, the other from the father. These chromosomes are the same in that they carry the same genes (e.g. the gene for the blood antigen that determines the blood group), but the form of the gene (allele) may not be the same – we could inherit the allele for blood group B from the mother and the allele for blood group A from the father. Before the introduction of DNA analysis in forensic genetics, the identification of blood groups played an important role in criminal investigations (Figure 15.1).

Thus, our DNA is a combination of the DNA of our ancestors, while the process of meiosis - during which sex cells are formed and recombination between homologous (same) chromosomes occurs - significantly contributes to the increase in genetic variability. Since the Y chromosome is found only in men, it is inherited through the paternal lineage. In addition to the DNA inside the nucleus of the cells, DNA is also found in the mitochondria, which are stored in the cytoplasm of the cell. After fertilisation, the zygote acquires the cytoplasm of the mother's egg cell, and therefore the mitochondrial DNA is inherited from the mother. With the help of mitochondrial DNA analysis, we can trace the maternal lineage. What is interesting here is that mitochondrial DNA is found in multiple copies, so even if the nuclear DNA present in the forensic trace were degraded, mitochondrial DNA may still be detectable.

Human genetic information consists of 3x109 base pairs. We refer to the order of individual nucleotides as the DNA sequence and the method by which we can determine this order is called sequencing (more in Chapter 4 - How do you work with DNA?). The first, almost complete sequence of human DNA was revealed in 2003, thanks to the Human genome project (HUGO). HUGO found, among other things, that the DNA of two unrelated people differs only in 0.1-0.3% of positions out of a total of three billion possible positions. In the case of identical twins, the DNA sequence is the same. Genes are the regions of DNA that code for some functional product (see Chapter 3 - Meet DNA, the bearer of genetic information). Genes make up only about 1.5% of the total genetic information of a person (it is assumed that a person has 20,000-22,000 protein-coding genes). The majority of genetic information consists of the so-called non-genic or non-coding DNA. Some of these non-coding regions have a regulatory function and can influence the transcription of genes, but in general, not much is known about the function of several parts of non-coding DNA.

If we want to compare the DNA of two people and distinguish them based on such an analysis, we have to focus on positions that are different between people. We refer to such positions as DNA polymorphisms. An important type of polymorphism is sequence polymorphisms, often referred to as single nucleotide polymorphisms - SNPs (Figure 15.2). This means that if, for example, an individual has an adenine at a certain location, someone else will have a guanine at that exact position in their DNA.

One of the methods by which we can determine differences in DNA between people is by restriction fragment length polymorphism analysis - RFLP. In this method, we use enzymes (restriction endonucleases) isolated from bacteria, which can specifically cleave (cut) DNA based on the sequence found in a certain place (Figure 15.3). For example, the EcoRI enzyme cleaves DNA only if it recognises consecutive GAATTC nucleotide sequence. If this sequence is the place of a DNA polymorphism, for example a cytosine instead of thymine in a given position, then the DNA can not be cleaved by the EcoRI enzyme at that place. This will then give a different pattern when run on a gel electrophoresis.

However, if we cut our entire DNA with such an enzyme, it would cut it in many places, which would greatly complicate the analysis of the results. Therefore, we have to define a section of DNA in which we know that the desired polymorphism occurs, and then analyse just that piece. We use the polymerase chain reaction (PCR) method to limit and multiply a certain defined section of DNA. This method was invented and described in 1983 by Kary Mullis and is explained in more detail in Chapter 4 - How do you work with DNA. Any cells can be used as a material for DNA isolation (e.g. lymphocytes, hair roots, sperm, skin cells...), and for profiling people, a swab of the buccal mucosa (inner side of the cheek) is most often used.