DNA Profiling and STR Typing in Forensics: From Molecular Techniques to Real-World Applications

Table Of Content
1. Foundations and Evolution of DNA Profiling
2. DNA Profiling Techniques
3. Classification of Forensic DNA Techniques
4. Short tandem repeats (STRs)
5. Steps in DNA Typing
6. STR Profiling
7. Real-World Applications of STR Typing

The Pioneers and Progress of DNA Profiling
Importance of DNA in forensics
DNA, the fundamental blueprint of life, has become one of the most powerful tools in modern
forensic science. From solving cold cases to exonerating the innocent, the journey of DNA profiling
has reshaped justice systems worldwide.
According to Evelyn Fox Keller (2000), the twentieth century was century gene, a period neatly
flanked by "rediscovery" of Mendel's principles of heredity at the beginning and the completion of the
first draft of the human genome at the end. During the second half of this epoch it became widely
accepted that DNA was the molecular basis of the gene.
DNA is famously described as a "double helix"-a molecule composed of two twisting strands. As a
carrier of genetic information, its key feature is the ordering of four chemical units called “bases"
known as adenine (A), thymine (T), cytosine (C) and guanine (G). The two strands of DNA's double
helix are held together through the complementary pairing of base A on one strand with T on the
other, and C on one strand with G on the other. This "complementarity" between pairs of bases
explains the faithful replication of the DNA molecule (and genetic information) from one generation
to the next. According to the principles of molecular genetics, the sequential ordering of these four
bases encodes genetic information, and differences in their arrangement are the basis of genetic
differences between species and between individuals of the same species. Each human cell contains
about two meters of DNA located in a compartment called the nucleus. Here it is tightly packaged into
twenty-three pairs of chromosomes, each of which contains a single DNA molecule of, on average,
roughly 150 million base pairs. The totality of nuclear DNA in a cell-which in most people is virtually
identical in almost every cell in the body-is popularly known as a "genome".
In 1992, Walter Gilbert, one of the early proponents of the Human Genome Project (HGP),
proclaimed that one day it would be possible to hold up a CD of one's own genomic DNA sequence
and say, “Here is a human being; it's me!" (Gilbert, 1992). The mapping of the concept of the gene
onto the biomolecular properties of DNA underpins one of the most persuasive arguments for
undertaking the HGP, namely, that the DNA genome is the "Book of Life" or, to modernize the
metaphor, "Life's CD." According to this metaphor, biology, destiny, and identity are encoded within
the molecular structure of the DNA genome.
DNA analysis is widely regarded as the "ultimate method of biological individualization" hence its
place in forensic science. However, there are two key differences between DNA analysis for forensics
and DNA analysis for the HGP. One is that forensic analysis does not examine the entire genome of
individuals, but a much smaller number of highly variable regions, comprising approximately one
millionth of a genome. Secondly, forensic science is not primarily interested in those parts of genomic
DNA that comprise genes. On the contrary, forensic analysis has favored DNA sequences that are
external to the genes, and are not believed to encode biological traits.
Approximately 99.5 percent of the human genome has been found to be the same for everyone.
Forensic techniques exploit those places in the genome, known as "polymorphic loci," which exhibit
detectable variations. These detectable variations (called “allelomorphs" or "alleles") can be of two
types: variation in the sequence of DNA bases; and length variation arising from differences in the
number of DNA bases between two defined end points.

Dr. Alec J. Jeffreys: legal application
'DNA fingerprinting' or DNA typing (profiling) as it is now known, was first described in 1985 by an
English geneticist named Alec Jeffreys. Dr. Jeffreys found that certain regions of DNA contained
DNA sequences that were repeated over and over again next to each other. He also discovered that the
number of repeated sections present in a sample could differ from individual to individual. By
developing a technique to examine the length variation of these DNA repeat sequences, Dr. Jeffreys
created the ability to perform human identity tests.
His method was first applied in the 1986 case of 15-year-old Dawn Ashworth’s rape and murder.
Although the prime suspect, Richard Buckland, had confessed, DNA analysis by Dr. Jeffreys revealed
that his DNA did not match samples from the crime scenes. This led to a mass DNA screening of over
4,000 local men. The real breakthrough came when Colin Pitchfork was identified after trying to
evade the screening using a stand-in. His DNA matched the crime scene samples, leading to his arrest
in September 1987 and conviction in early 1988—the first murder conviction secured using DNA
evidence. That same year, DNA evidence was also used to convict Tommy Lee Andrews for rape in
the U.S.
DNA FINGERPRINTING
DNA fingerprinting technique began years ago with the introduction of restriction fragment length
polymorphism (RFLP), and in the 1990s, RFLP method gave way to PCR methodologies that had the
advantage of being able to amplify DNA. After several improvements and refinements to the PCR-
based tests, the forensic community came to an agreement on the use of STRs. Although there are
other DNA markers in use, STR typing is the method of choice for most forensic laboratories, and
given the investments in infrastructure, training, databases, and accreditation, it will be for the
foreseeable future. Evidentially, a wide range of microsatellite loci have been identified,
characterized, and demonstrated to be highly abundant in the human genome. The high abundance and
polymorphic nature of STR loci was a vital factor considered for incorporation into commercial kits
by manufacturers, and has ever since then remained the most frequently applied methodology, and the
current gold standard for human identification in forensic laboratories.

Types of forensic DNA analysis:
Forensic DNA analysis has come a long way since its early beginnings in the 1980s, with the
emergence of several different techniques. STR analysis is now the primary method but, before its
invention, restriction fragment length polymorphism (RFLP) was the dominant technique.
Additionally, since the adoption of STR approaches, other methods such as Y chromosome analysis,
mitochondrial DNA (mtDNA) analysis, single nucleotide polymorphism (SNP) typing and mini STR
analysis have also been developed. This chapter explores the evolution of forensic DNA analysis
techniques, and explains when to use which alternative to STR typing.
RFLP System:
The first forensic DNA typing systems were based on a kind of length variation called "restriction
fragment length polymorphism" (RFLP). "Restriction fragments" are produced by treating a DNA
molecule with a "restriction enzyme," which acts like a pair of "molecular scissors," making a precise
cut every time it finds a specific short sequence of bases which are distributed throughout the DNA.
Restrictions fragment length polymorphism, initially established in the middle of the 1980s, was the
first DNA typing method. DNA was typed using the RFLP method, which comprised central elements
of sequences made up of 30-100 repetitions (variable number tandem repeats). A significant amount
of intact genomic DNA is needed for DNA characterization using the restriction fragment length
polymorphism technique (20 to 30 mg). However, the biological samples brought into a lab for
forensic science are frequently subjected to environmental abuse, and sometimes only trace amounts
of DNA may be extracted. As a result, the restriction fragment length polymorphism technique was
often inapplicable.
VNTRs:
"VNTR profiling" uses restriction enzymes to identify highly polymorphic loci in variable number
tandem repeat (VNTR) regions. A typical VNTR locus is composed of a short sequence of DNA bases
(the "repeat unit") which is repeated a variable number of times.4 Alleles at a VNTR locus differ from
one another in length depending on how many repeat units they contain. A restriction enzyme that cuts
DNA on either side of a VNTR will generate restriction fragments of lengths that are dependent on
which VNTR alleles it contains. To detect these allelic differences, the fragmented DNA is loaded
onto an indentation at one end of a flat plate made of agarose gel, and an electric current is applied
across the gel. Under the influence of the electric field, the DNA fragments are pulled out of the well
and through the jelly-like substance in a straight line (called a "lane"). The smaller the fragments, the
DNA is Extracted Restriction Digestion
Electrophoresis separates the DNA fragments

faster they travel. at the end of the "electrophoresis” process the fragments are separated from each
other and arranged in length order along the lane.
Fragments containing loci of interest are picked out from all the other DNA fragments using "probes"
labeled with a radioactive tag. Each probe is a short sequence of DNA that is "complementary" to the
sequence in a VNTR unit. The probe seeks out and "hybridizes" with (i.e., binds to) all the VNTR
fragments on the membrane that contain the complementary sequence. The gel is then pressed against
a nylon membrane to transfer the ordered DNA fragments onto the membrane for analysis. When the
image ("autoradiograph") of the membrane is developed, each bound probe appears as a band whose
position reflects the size of the allele on the fragment.
PCR Systems:
PCR was invented in 1986 by Kary Mullis at Cetus Corporation in California. Sometimes referred to
as "molecular Xeroxing," the PCR technique uses an enzyme called a "polymerase" to replicate (or
"amplify") a DNA sequence 100-2,000 base pairs long. PCR produces millions of copies of the initial
DNA sequence through a chain reaction in which the products of one round of replication become
templates for the next. In each round, the precise DNA sequence replicated is defined by two synthetic
"primers" (special probes) that bind to either end, one to initiate the replication reaction and the other
to terminate it. A PCR system can generate profiles from samples as small as 0.3-0.5 nanograms of
DNA, about a hundred times smaller than required for RFLP, and corresponding to the amount of
DNA present in a few hundred sperm cells or a blood spot the size of a large pin- head.

AFLPS:
"Amplified fragment length polymorphisms" (AFLPs) are DNA fragments of differing lengths that are
generated by amplifying a polymorphic DNA region with PCR. For a brief period, a few forensic
DNA laboratories used AFLPS of a VNTR locus called DIS80. In this system, PCR was used to
generate millions of copies of DIS80 alleles in the sample. These were then sorted by size using
electrophoresis and visualized as a profile of one or two bands representing each person's genotype at
this locus.
COMPARISON OF DNA TYPING METHODS
Forensic DNA markers are arbitrarily
plotted in relationship to four quadrants
defined by the power of discrimination
for the genetic system used and the
speed at which the analysis for that
marker may be performed. Note that
this diagram does not reflect the
usefulness of these markers in terms
of forensic cases.
Short tandem repeats (STRs)
also known as microsatellites or simple sequence repeats, are short tandemly repeated DNA sequences
that involve a repetitive unit of 1-6 bp, forming series with lengths of up to 100 nucleotides. STRs are
widely found in prokaryotes and eukaryotes, including humans. They appear scattered more or less
evenly throughout the human genome, accounting for about 3% of the entire genome. However, their
distribution within chromosomes is not quite uniform—they appear less frequently in subtelomeric
regions. Most STRs are found in the noncoding regions, while only about 8% locate in the coding
regions. Moreover, their densities vary slightly among chromosomes. In humans, chromosome 19 has
the highest density of STRs. On average, one STR occurs per 2,000 bp in the human genome. The
most common STRs in humans are A-rich units: A, AC, AAAN, AAN, and AG.
Eukaryotic genomes have repetitive DNA sequences. These DNA sequences are very small in size and
are usually determined by the length of the nucleus of each repetitive unit and the number of repeats
of the nucleus of each unit. Regions of DNA have repetitive units’ 2 to 7 bp microsatellite, or
commonly called short tandem repeats (STRs).

Types of STR Markers
On the basis of different repeat units, STRs can be classified into different types. On the one hand,
according to the length of the major repeat unit, STRs are classified into mono-, di-, tri-, tetra-, penta-,
and hexanucleotide repeats. The total number of each type decreases as the size of the repeat unit
increases. The most common STRs in the human genome are dinucleotide repeats.
On the other hand, according to the repeat structure, STRs are classified into:
• Perfect repeats (simple repeats), containing only one repetitive unit
• Imperfect repeats (compound repeats), consisting of different composition repeats.
The STR locus is named as, for example, D3S1266, where:
D DNA
3 Chromosome 3
S STR
1266 Unique Identifier
Classification of tandem repeats:
I. STRs (Short tandem repeats or micro-satellites): Short tandem repeats are sequences where the
repeating unit has a base pair length of 1-6 bps.
II. VNTRs (Variable number tandem repeats or mini-satellites): Variable number tandem repeats are
sequences where the repeating unit has a length of 7 to 100 bps (base pairs).
III. Satellite DNA: The sequence is referred to be satellite DNA if the repeating unit is between 100
and several thousand base pairs in length.
A common nomenclature for STR alleles
A repeat sequence is named by the structure (base composition) of the core repeat unit and the number
of repeat units. However, because DNA has two strands, which may be used to designate the repeat
unit for a particular STR marker, more than one choice is available and confusion can arise without a
standard format.
For example, to produce a ladder containing five alleles with 6, 7, 8, 9, and 10 repeats, individual
samples with genotypes of (6,8), (7,10), and (9,9) could be combined. Alternatively, the combination
of genotypes could be (6,9), (7,8), and (10,10) or (6,6), (7,7), (8,8), (9,9), and (10,10).
THE 13 CODIS STR LOCI
In the United States, utilization of STRs initially lagged behind that of Europe, especially the efforts
of the Forensic Science Service in the United Kingdom. However, beginning in 1996, the FBI
Laboratory sponsored a community-wide forensic science effort to establish core STR loci for
inclusion within the national DNA database known as CODIS (Combined DNA Index System).
Chapter 18 covers CODIS and DNA databases in more detail. This STR Project beginning in April
1996 and concluding in November 1997 involved 22 DNA typing laboratories and the evaluation of

17 candidate STR loci. The evaluated STR loci were CSFIPO, F13A01, F13B, FES/FPS, FGA, LPL,
TH01, TPOX, VWA, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, and
D21S11.
At the STR Project meeting on 13-14 November 1997, 13 core STR loci were chosen to be the basis
of the future CODIS national DNA database. The 13 CODIS core loci are CSFIPO, FGA, TH01,
TPOX, VWA, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, and D21S11.
When all 13 CODIS core loci are tested, the average random match probability is rarer than one in a
trillion among unrelated individuals.
The three most polymorphic markers are FGA, D18851, and D21S11, while TPOX shows the least
variation between individuals.
Using the previously described classification scheme for categorizing STR repeat motifs, the 13
CODIS core STR loci may be divided up into four categories:
1. Simple repeats consisting of one repeating sequence: TPOX, CSFIPO, D5S818, D13S317,
D16S539;
2. Simple repeats with non-consensus alleles: TH01, D18S51, D7S820;
3. Compound repeats with non-consensus alleles: VWA, FGA, D3S1358, D8S1179;
4. Complex repeats: D21S11.
Y-Chromosome STRS
The "sex" chromosomes X and Y contain STR loci that can be PCR- amplified. Because women are
assumed not to have a Y chromosome, Y-STR profiling is used to distinguish between male and
female “donors" of mixed crime stains, for example in sexual assault cases.
The use of Y STRs can be effective in the study of mixed biological specimens (e.g. in cases of rape).
In cases where the DNA of a male is mixed with the high level of DNA of a female; although there
must be a specific sexual incompatibility for the use of Y-STRs. However, Y-STR analysis is of great
help in identifying DNA specimens with a female DNA background. However, depending on the
frequency of haplotype Y in the population, its statistical value can be limited, and the possibility of
the presence of DNA of the relatives of a suspect’s paternal lineage cannot be ruled out as a
contributor in the stain.
DNA typing process
The technique begins with a reference sample, which is a DNA sample from a human. The ideal
method for acquiring a reference specimen is to use a buccal swab since it decreases the occurrence of
infection. Alternative approaches may need to be used to get a specimen of blood, saliva, semen, or
another acceptable fluid or tissue from a person or previously kept samples if this is not possible for
example, if a court order is necessary but not always attainable) (e.g., banked sperm or biopsy tissue).
The analysis of a reference specimen is then used to generate the person's DNA profile. The DNA
profile is then contrasted with a separate specimen to see whether there is a genomic equivalence.
Figure shows the steps involved in DNA fingerprinting technique.

STR Typing
Years ago, restricted fragment length polymorphism, a DNA profiling technique, was abandoned in
favour of PCR procedures, which amplified DNA. After numerous PCR-based procedures were
improved upon and modified, STRs were accepted by the forensic community. Due to the expense of
infrastructure, training, databases, and certification, STR typing will continue to be the technique of
selection for most forensic laboratories, even if additional DNA markers are utilized. Microsatellite
loci have been discovered, recognized, and established to be remarkably prevalent in the human
genetics. When commercial kits were created, the enormous abundance and polymorphism
characteristics of STR loci were taken into consideration. Since then, forensic laboratories have
continued to employ this method as the most popular and current industry standard for using human
identification.
Steps for STR Analysis
In order to perform analysis on STR markers, the invariant flanking regions surrounding the repeats
must be determined. Once the flanking sequences are known then PCR primers can be designed and
the repeat region amplified for analysis. New STR markers are usually identified in one of two ways:
(1) searching DNA sequence databases such as GenBank for regions with more than six or so
contiguous repeat units or (2) performing molecular biology isolation methods.
The recognized DNA typing approach is followed by STR typing analysis. On the other hand, the
STR typing method is based on the standard operating procedures (SOPs) supplied by the
manufacturers of the commercial kit for use in forensic laboratories. The following steps are taken for
STR typing in the conventional order for DNA typing:
• DNA extraction is performed to decide the amount of DNA present
• amplify STR loci
• separate PCR amplicons on a genetic analyzer
• examine the resultant data using bioinformatics
• and compare the data from one sample to directories with previously generated short tandem
repeats sets.
The size and colour of these amplicons may be used to differentiate STR loci, which are repetitive
DNA sequences with variable repetition counts produced using primers with distinct fluorophores.
Another crucial step in analyzing STR loci is identifying the invariant flanking areas surrounding the
repeats. If the adjacent sequences are known, the repeat region may be amplified using PCR primers.
Typically, there are two methods to calculate short tandem repeats. These comprise using molecular

science isolation techniques or searching DNA sequence directories like Combined DNA Indexed
System or GenBank for areas above vicinal repeat units.
STRs are highly polymorphic and alleles of the STR loci are differentiated by the number of copies of
the repeat sequence within each of the STR locus. Research findings have demonstrated that the more
STR loci being used for typing, greater the discrimination value, since the probability that two
individuals taken from a random population possessing exactly the same number of repeats units for
all the STR being analyzed, is extremely rare. They can vary in size from person to person without
impacting the genetic health of the individual. For example, at the same locus, a tetra-nucleotide
repeat sequence (represented by CTAG) will vary from one person to the other. As in the figure,
Person 1 has 5 repeats, person 2 has 6 repeats and person 3 has 7 repeats.
Forensic DNA profiling:
• Targeting Specific DNA Regions with Primers
In the simple model provided so far, all DNA would be replicated with no start and end points on the
DNA strand. To identify the specific areas of interest (the "target"), it is necessary to identify specific
sequences of DNA on either side of the target STR that will enable only that DNA to be amplified.
Primers are short lengths of DNA that will bind specifically to the areas on either side of the target
locus. Two primers are required for each locus.
• Separation by Capillary Zone Electrophoresis (CZE)
The product of the amplification process is a soup of DNA molecules of different lengths, which are
the alleles at each locus. One way to visualize the components of any mixture is simply to separate the
components. This is achieved in routine DNA profiling by a technique called capillary zone
electrophoresis (CZE).
• Amplification and Allele Generation
During the process of amplification, the DNA molecules are "tagged" with dyes of different colors.
These enable the detection of the DNA by detection of the colored light (in reality, the dyes are
illuminated by a laser and the light emitted by the dyes is then detected).

• Detection and Analysis Using EPG
In the separation process, charged DNA molecules are pulled through a narrow tube called a column
by an applied voltage. Smaller molecules move faster and reach the end sooner than larger ones. As
molecules exit the column, they are detected and converted into an electronic signal, which is
processed by computer software into a graph called an EPG. The analyst examines the EPG to
identify the alleles present in the sample.
An EPG, from what was the standard kit used in the United Kingdom for many years (SGM+,
manufactured by Applied Biosystems), is shown in Figure. Other kits may have different loci,
different numbers of loci, and different numbers of colored channels, but produce similar epgs.
It is probably easier to examine a single piece of a profile in isolation to get to know the various parts.
In Figure 2 (expanded), the loci are identified by the gray boxes above the three "channels" shown on
the epg (locus label). Every graph has two axes, the vertical (y-axis) and the horizontal (x-axis). In the
epg, the horizontal axis generally shows the size of the DNA molecule, which appears; smaller
molecules will show as peaks toward the left-hand side of the epg and larger molecules to the right.
Technically, the set of forensic STR loci is analyzed using multiplex PCR, and the amplified
fragments are sized using capillary electrophoresis (CE). One of the two primers amplifying each
locus is labeled with a fluorophore, allowing for detection. Four (or five) different fluorophores are
assigned to the various loci in such a way that fragments of each STR locus can be unequivocally
identified based on size and color on the electropherogram. A heterozygous genotype of a particular
locus thus will display two peaks of similar height on the electropherogram, whereas a homozygous
genotype will display one peak.
Schematic depiction of an
electropherogram of an STR
analysis showing the common
stochastic effects of low template
(LT) DNA (bottom panel) as
compared to optimal DNA
amounts (upper panel) from the
same individual. The STR loci
are indicated in italics, and the
blue rectangles encompass the
size ranges of amplicons of the

particular STR loci. Thus, peaks within that range are assigned to the respective STR loci, and
numbers in rectangles indicate the allele numbers of the peaks. Asterisks indicate stutter peaks. ADO:
allele drop-out; ADI. Allele drop-in. The fragment length in base pairs (bp) is indicated on the x-axis
on top, and the y-axis represents the peak height in relative fluorescence units (RFU). Please note the
different scale in RFUs between the two panels.
Example for STR data Interpretation using an EPG
Comparison of the DNA profiles for two
individuals obtained with multiple short
tandem repeat markers. STR length variation at
unique sites on 10 different chromosomes are
probed with this DNA test to provide a random
match probability of approximately 1 in 3
trillion. A gender identification test also
indicates that the top sample is from a male
while the bottom sample is from a female
individual. These results were obtained from a
spot of blood the size of a pin head in less than
five hours. The DNA size range in base pairs is
shown across the top of the plot. Results from
each DNA marker are indicated by the letters A-J.
Stutter Peaks
One important type of technical artifacts that typically occur in STR analysis are so-called stutter
peaks, which result from the propensity of the repeat units to slip by one or more units during the
elongation step of PCR amplification. As a consequence, stutter peaks are seen as small peaks
preceding the main peaks and are typically one complete repeat unit shorter than the true alleles.
Stutter peaks with sizes that are one unit longer or two or more repeat units different from the main
peak sometimes occur as well. The incidence of replication slippage in a particular PCR assay is
characteristic for each locus, and thus either a general stutter threshold or locus-specific stutter
thresholds are applied. Preclusion of stutter peaks is important because they have the same lengths as
expected for true alleles and would lead to wrong interpretations of electropherograms.
Principle of allelic ladder formation
STR alleles from a number of samples are separated on a polyacrylamide gel and compared to one
another. Samples representing the common alleles for the locus are combined and re-amplified to
generate an allelic ladder. Each allele in the allelic ladder is sequenced since it serves as the reference
material for STR genotyping. Allelic ladders are included in commercially available STR kits.

Practical Uses of STR Profiling in Forensics and Beyond
Characterizing Cell Line:
STR typing with the same core set of markers and commercial STR kits is being used for several
other purposes besides forensic and parentage testing.
Human cell line authentication is now being carried out by the American Type Culture Collection
(ATCC) along with other international suppliers of cell lines. STR typing enables rapid discovery of
cross-contamination between cell lines and may serve as a universal reference standard for
characterizing human cell lines. This type of analysis has been dubbed 'cell culture forensics'. Over
the past several years, the ATCC has created a database of over 500 human cell lines that have been
run with eight STR loci present in the PowerPlex® 1.2 kit from the Promega Corporation. It is
important to note that cell cultures, such as K562. may not always have a regular diploid complement
of chromosomes and thus may possess tri-allelic patterns and severe peak imbalances due to the
presence of additional copies of one allele.
Monitoring Transplants:
Monitoring the engraftment of donor cells after bone marrow transplants or allogeneic blood stem cell
transplantation is another important application of STR testing. Examination of STR profiles from
transplant recipients can help diagnosis graft failure or a relapse of the disease. In these cases,
mixtures are detected as mixed chimerism that exists within the recipient from their own cells and
those of the donor.
Detecting Genetic Chimeras:
Chimerism, which is the presence of two genetically distinct cell lines in an organism, can be acquired
through blood stem cell transplantation, blood trans- fusion, or can be inherited. Approximately 8% of
non-identical twins can have chimeric blood. Several years ago an interesting case was reported of a
phenotypically normal woman who possessed different DNA types in different body tissues due to
tetragametic chimerism.
A study with 203 matched related donor-recipient pairs ranked 27 different STRS, including the 13
CODIS core loci, in terms of their ability to detect chimeric mixtures. Not surprisingly, the loci with
the highest heterozygosities, namely Penta E, SE33, D2S1338, and D18851, worked the best.
Monitoring Needle Sharing:
In yet another application of the capability to perform mixture detection with STRS, a laboratory
method was used using the CODIS STR marker D8S1179 to differentiate between single and multi-
person use of syringes by intravenous drug users. Monitoring needle sharing can help determine the
source of spreading blood-borne pathogens among drug users.
Detecting Cancer Tumors:
Loss of heterozygosity (LOH) is a method of monitoring genetic deletions common in tumors for
many types of cancer. LOH is manifested by severe allelic imbalance at a locus in a single-source
DNA sample so that a true heterozygote almost appears as a homozygote since some of the
chromosomes have a deletion present in the region of the locus being PCR-amplified.

Probably the only time that LOH would have an impact on human identity testing is if an archived
clinical specimen from a tissue biopsy was used as a reference sample to identify someone from a
mass disaster. However, it is worth being aware of the fact that normal and cancerous tissue from an
individual can vary fairly dramatically in some instances in terms of their STR allele peak heights. An
examination of a cancer biopsy tissue specimen compared to normal tissue with the nine STR loci
present in the AmpF/STR Profiler kit found that the D13S317 locus exhibited a severe peak imbalance
consistent with that seen arising from LOH. The authors suggest that this LOH might be due to a
deletion of 13q21-22 seen previously with prostate cancer that is near the physical location of
D13S317 on chromosome 13.
Examining Human Population Diversity:
Whole genome scans with 377 autosomal STR loci were used to genotype 1056 individuals from 52
populations in order to study human population structure. Studies with this same set of data have
identified particular STR loci that are effective indicators of ancestral origin. Analysis of Y
chromosome STRS and mitochondrial DNA have also been used for genetic genealogy studies. Both
STR markers and single nucleotide polymorphisms (SNPs), which will be discussed in the next
chapter, should continue to play an important role in understanding human diversity at the genetic
level.
Analysis of the Y Chromosome:
Typically, biologically a male individual has 1 Y chromosome and contains 55 genes. Because of this
unique feature, analysis of Y chromosome is done in crime cases.
Application of Y chromosome in forensic medicine: It is present only in males. Thus, in crime
cases, the investigators expect to find Y chromosome at the crime scene. Also, when talking about
male–female ratio in body fluid mixtures, such as sexual assault or rapes, by analyzing the Y-STR
component, the investigators can obtain more information regarding the male component. It is well
known that azospermic or vasectomized rapists do not leave semen traces, and it is impossible to find
spermatozoa on the microscopic examination. In such cases, the Y-STR profiling is very useful,
offering information regarding the identity of the accused person.
Impact of Genetic Identification in Justice
Genetic testing using DNA has been widely applicable to the field of justice. This method is being
used for the following:
• Identification of accused and confirmation of guilt.
• Exculpation of innocent ones.
• Identification of persons who commit crimes or serial killers.
• Identification of victims in disasters.
• Establishing cognation in complex cases.

Key Properties:
STRs have such properties as abundant, codominant, highly polymorphic, and nearly selectively
neutral. Besides, STRs contain DNA fragments that are small enough to be amplified by polymerase
chain reaction and separated in high-resolution media like polyacrylamide. With the availability of
high-throughout capillary sequencers or mass spectrography, the sizing of alleles is no longer a
bottleneck in STR analysis. Thus STRs are widely used in scientific and applied research.
STRs in Genetics and Population Studies:
STRs are extremely useful in applications such as the construction of genetic maps, gene location,
genetic linkage analysis, identification of individuals, paternity testing, as well as disease diagnosis.
STR analysis has also been employed in population genetics. Nevertheless, the application of STRs to
population genetics requires a more detailed understanding of the STR mutation process.
in Evolutionary Studies:
We can apply STRs to reconstruct the history of migration and evolution of the species, as well as to
assess biological diversity at various levels of biological organization. A method of absolute genetic
dating uses mutation rates as molecular clocks. Such a molecular clock based on STR, whose
mutation rate is very high, can be applied to human evolution. Therefore, STRs are likely to reflect
relatively recent divergence.
Phylogenetic Insights and Mutation Dynamics:
The difference in size between two different STR alleles might be informative: the larger the
difference, the more the number of mutation events. Thus there is a “memory” of past mutation
events. That is, when a mutation occurs, the new mutant is related to the allele from which it was
derived. In this case, the difference in length between alleles contains phylogenetic information.
However, the prevalence of different mutational events may vary dramatically among groups.
Ignoring the possibility that the same allelic type found in different individuals or populations may be
derived from different evolutionary processes, it might lead to biased estimates of genetic structure.
Consequently, it is very important to know the mutation process of STRs in detail before they are
applied to population genetics studies.
Mutation Models:
Mutation models for the evolutionary process of STRs are needed in order to estimate phylogenetic
relationships, population differentiation measures, and genetic distances from STR data. Different
kinds of estimators based on IAM have been developed, such as DAS (shared allele distance), DCH
(Cavalli-Sforza and Edwards chord distance), and DS (Nei’s standard genetic distance). On
SMM/TPM, estimators include (δμ)2, DSW (stepwise weighted genetic distance), and RST. Different
estimators can be effective in different situations. It was concluded that for a relatively short period of
time, DAS or DS is a better measure, but as time increases, the estimator based on SMM such as
(δμ)2 becomes superior.
in Tracing Human Evolution:
In 1995, Goldstein et al. predicted that STR loci would ultimately allow a high-resolution description
of the human evolutionary history. Many researchers have studied the history of human evolution and
migration by using STR loci. Scientists developed a new combination polymorphism, namely

SNPSTRs, in which each such segment includes one or more single nucleotide polymorphisms
(SNPs) and exactly one STR locus, providing insights into population history. At present, STR loci
are employed to reveal the relationship of populations in different regions, as well as the route of
migration of ancient peoples.
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DNA Profiling and STR Typing in Forensics: From Molecular Techniques to Real-World Applications

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DNA Profiling and STR Typing in Forensics: From Molecular Techniques to Real-World Applications