Information Available From nDNA

Nuclear DNA (nDNA) is used to generate individual genetic profiles. A genetic profile is the result of gametes that have undergone random recombination. Each gamete has a unique selection of DNA from each parent, has an equal opportunity to be selected, and will produce a zygote that is unique. Only identical twins have identical DNA. The same is true of animals. The sex of an animal is determined by identifying the XY chromosomes in mammals (females are XX and males are XY) or the WZ chromosomes in avians (females are WZ and males are WW). Some species such as crocodilians, turtles, and tortoises have temperature-dependent sex determination, which means they must have a full complement of both sets of sex chromosomes so they cannot be genetically sexed. However, individual identification is still possible.

nDNA is the gold standard for identifying individuals. In a criminal case, the evidence needs to be processed for DNA residue before the exemplars, the known DNA contributors, are processed. Evidence may be blood stains or saliva collected from a fighting pit, bites, blood in the back of a truck, on the ground, hairs from kennels or crates, teeth, bones, or clothing. After the species has been established using mtDNA, the gender of the donor (X/Y chromosomes from nDNA) and the unique profile are determined from nDNA. A genetic profile can only be generated if the markers are available. If markers and a database are available, then the genetic profile can be compared to the profiles generated from the exemplars, such as buccal swabs taken from dogs, tissue samples from dead deer or other livestock, or perhaps hair samples taken from a captured bear. The information obtained will help to determine if the dogs were in a specific fighting pit, which dogs were involved in a biting or mauling incident, which bear was involved in a home invasion, or if a doe was taken out of season. DNA evidence can immeasurably strengthen a case.

A genetic profile is generated by using multiple markers that take advantage of a particular class of repetitive DNA, referred to as short tandem repeats (STRs). These microsatellites repeats are 2 to 6 nucleotides that are repeated usually 8 to 20 or more times, like a stutter. The number of repeats at a homologous site often differ between the mother and the father. This is nDNA, so one strand of DNA comes from the mother and the other strand from the father. They may, by chance, have the same number of repeats or a different number of repeats. The combination of differences at multiple sites becomes a genetic profile that is unique for each individual. The process used to assay the loci, PCR, allows for the lengths of both alleles (one from each parent) at each locus to be identified and visualized (Figure 4.4).

Processing a crime scene and collecting evidence is dealt with in detail in Chapter 1. It is important to remember that when evidence is being collected for DNA analysis, gloves must be worn, wet samples allowed to dry if at all possible, and items must be properly packaged to prevent contact contamination or contamination from investigators. All samples must be properly labeled with date, location, case number, collector, an individual identifier, and a brief description. The laboratory technician was not at the scene and will not have any reference for a sample except for the identifier and description on the container.

(a) Examples of STRs

Target region (short tandem repeat)

TCTA TCTA ; TCTA TCTA TCTA | TCTA TCTA : TCTA [—

= TCTA TCTAl TCTA TCTA,TCTA |TCTA,TCTA ! TCTA TCTA TCTA allele 10

DNA Profile of this locus = 8,10

PATERNITY TESTING

Family Inheritance of STR Alleles (Dl 3S317) PCR produrt size (bp)

ISO 190 200 210 220 230 240 250

Figure 4.4 Visualization of STRs (short tandem repeats) (a) STRs are made up of short repeats of DNA. One strand (allele) is from the mother and the other strand from the father. They may be the same number or a different number of repeats, (b) This is an illustration of peaks at one locus used for analysis. The father and mother are heterozygous (two different-sized peaks: 11/14 and 8/12, respectively) at this one locus. Child 1 (12/14) is also heterozygous, getting the 12 allele from the mother and the 14 allele from the father. Child 2 (8/14) received the 8 allele from the mother and the 14 allele from the father. Child 3 (11/12) received the 11 allele from the father and 12 allele from the mother. This is one locus. Many loci added together result in a profile that is unique to the individual, ([a] From: https://www.slideshare.net/RanaMuhammadAsif/forensic-dna-typingm-asif-59208013. [b] https://slideplayer.com/slide/4798394/.)

It is good to over-collect evidence samples. The scene is secure only once and it is difficult to go back for more evidence collection. Not all samples need to be processed, only those necessary to address the questions of the case. Also, if a sample fails, then there is more material to go back to for analysis.

Evidence needs to be stored appropriately, and the conditions when collected should be relayed to the laboratory technician, such as if there has been exposure to the elements, sun, or chemical compounds. All of these conditions accelerate the degradation of DNA, as does time. If evidence has been exposed to negative conditions, the technician will want to process the evidence in a timely manner to avoid more degradation and to capture the most information possible. mtDNA is hardier and less susceptible to degradation because it is circular and there is much more of it in each cell (10-1000 copies in each cell as compared to a single structure in the nucleus). nDNA begins to degrade as soon as there is cell death and the nucleus walls begin to break down. The sooner the DNA can be captured and the degrading process halted, the better the information that can potentially be extracted. To slow the degradation process once the DNA evidence has been collected, the evidence should be dried if collected wet. If it cannot be dried at the collection site, it should be frozen as soon as possible. These actions stop the enzyme and bacterial activity that initiate DNA breakdown. Samples that are collected in a warm, humid environment should be stored at the collection site in a cooler with ice packs until transportation to the final storage facility. This will protect the DNA evidence from further damage due to heat, exposure to sun, and sweating within its collection container. It is best to store evidence in paper or cardboard to prevent sweating. Plastic should be used only if there is risk of contact contamination or when the item is thoroughly dry, such as bones.