JIAOMR

10.5005/jp-journals-10011-1233
REVIEW ARTICLE Understanding the Basics of DNA Fingerprinting in Forensic Science

Understanding the Basics of DNA

Fingerprinting in Forensic Science
1
Vasavi Krishnamurthy, 2Reema Manoj, 3SS Pagare
1
Professor, Department of Oral Medicine and Radiology, Padmashree Dr DY Patil Dental College and Hospital
Navi Mumbai, Maharashtra, India
2
Lecturer, Department of Oral Medicine and Radiology, Padmashree Dr DY Patil Dental College and Hospital
Navi Mumbai, Maharashtra, India
3
Professor and Head, Department of Oral Medicine and Radiology, Padmashree Dr DY Patil Dental College and Hospital
Navi Mumbai, Maharashtra, India

Correspondence: Vasavi Krishnamurthy, Professor, Department of Oral Medicine and Radiology, V701, Guru Samridhi Height
Plot No. 2, Sector-14, Sanpada, Navi Mumbai-400705, Maharashtra, India, e-mail: sanvas72@yahoo.com

ABSTRACT

Scientific technology has recently made significant progress in the area of human genetics. The discovery of deoxyribonucleic acid (DNA)
has proven to be momentous for science. Dr Alec Jeffreys found that certain regions of DNA contained DNA sequences that were separated
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 which is the basis for its use in forensic science. Though the use of such knowledge is not fully realized yet, there is
limitless potential for the subject. This paper aims to describe the basics to understand the various methods of DNA fingerprinting and its
applications in forensic science.
Keywords: DNA fingerprinting, Forensic science, Genetic fingerprinting.

INTRODUCTION structures called chromosomes. In addition to DNA,

The study of human genetics has grown exponentially in recent
years, and from this scientific field and its accompanying
techniques scientists have been able to determine the difference
between individuals using DNA. Following discoveries of Dr
Alec Jeffreys, a British geneticist, a powerful new tool has been
developed for use in forensic investigations. Popularly known
as ‘DNA fingerprinting’ it is more accurately described as the
determination of the sequence of bonds on part of the DNA
strand within chromosomes, which is genetically determined
and capable of being related to other members of the same
family. Only uniovular twins have the same sequences;
otherwise the chances of two unrelated individuals sharing the
same sequence was one in a million billion and even amongst
siblings, only one in 10,000 million. Unlike blood groups, the
specificity of the DNA profiles is so great in statistical terms,
that it is greater than the human population of the world and is
reliable and specific to any individual.1
By developing a technique to examine the length variation
of these DNA repeat sequences Dr Jeffreys created the abiliy
to perform human identity tests. It was then called as genetic
fingerprinting.
First use of DNA testing in forensic setting was done in
1986. Understanding this new technology and its correct
implementation are keys for its acceptance throughout society.2
chromosomes contain associated proteins, predominantly
histones that organize the DNA into its native confirmation. In
most human cells, there are 46 chromosomes-one pair of each
type. Most cells have two sets of chromosomes and are said to
be diploid only excepting the male and female sex cells
(gametes) which are haploid and contain only one chromosome
of each type. The fertilized egg, the zygote, possesses 46
chromosomes-23 maternally derived and 23 paternally derived.3
The various combinations of the nucleotides adenine (A),
guanine (G), cytosine (C) or thymine (T) yield the diverse
biological differences among human beings and all living
creatures. Humans have approximately 3 billion nucleotide
positions in their genomic DNA.4
NATURE OF GENE3
The human genome contains approximately 3 billion base pairs
(bp) of DNA coding for an estimated 50,000 to 100000 genes.

DNA AND ITS STRUCTURE (FIG. 1)

DNA, the hereditary material of living organisms is primarily
found in the cell nucleus. DNA is organized into long threadlike Fig. 1: DNA structure

Journal of Indian Academy of Oral Medicine and Radiology, October-December 2011;23(4):613-616 613
Vasavi Krishnamurthy et al
Genes are situated in discrete regions, called genetic loci, on
the 46 chromosomes. In addition, most genes exist in multiple
forms called alleles. A single genetic locus may have many
different alleles and this variation is termed as genetic
polymorphism. A population may have multiple alleles at a
given locus and this genetic polymorphism is the molecular
basis of forensic DNA typing.
A vast majority of our DNA molecules (over 99.7%) is the
same between people. Only a small fraction on our DNA (0.3%
or 10 million nucleotides) differ between people and make us
unique individuals. These variable regions of DNA provide the
capability of using DNA information for human identity
purposes.2
GENETIC POLYMORPHISM AND
FORENSIC DNA TYPING
Investigators have identified >2500 polymorphic loci in the
human genome. Two main types of DNA polymorphism are
studied in forensic science: Single base changes or single
nucleotide polymorphisms (SNP) and minisatellite sequences
with VNTRs (variable number of tandem repeats). Both classes
of polymorphism are often recognizable as changes in DNA
fragment length following restriction enzyme digestion.5
Single nucleotide polymorphisms (SNPs) are differences
in the DNA sequence that occur when a single base in the
genome is altered. Single base polymorphism is caused by
deletion, insertion or substitution of one nucleotide. To further
elaborate, a SNP might change the DNA sequence
…AAGGCTAA to ….ATGGCTAA.
Since, the number of alleles at these loci is small, they have
limited discriminating power for forensic DNA identification.6
VARIABLE NUMBER OF
TANDEM REPEATS (VNTRs)
The class of polymorphic loci, known as VNTRs are highly
polymorphic in humans. Several characteristics of VNTRs—
the large number of alleles, their high heterozygosities and their
frequent occurrence in the genome make them highly
informative for purposes of human gene mapping, identification
of individuals and determining the biological relatedness
between individuals.7
They are also called as length polymorphism. For example:
………. (AATG) (AATG) (AATG) …………….. 3 repeats
………… (AATG) (AATG) ………………. 2 repeats.
Several well-characterized genetic loci contain core
elements (sets of nucleotides) that are tandemly repeated. The
number of these repeat units varies among individuals and size
of the core repeat varies between the loci.5 The variability of
this information makes each set of genetic information unique.
Polymorphisms of 9 to 80 bp (base pair) repeats are known as
VNTR.8 The advantage of examining specimens of VNTR is
the stability across generations. They also determine unique
sets of genetic markers for individual identification.
To date, over a hundred such VNTR or hypervariable
minisatellite (HVR) loci have been described and characterized
614
in the human genome. However, population-specific VNTR
allele frequency data form an integral part for the appropriate
utilization of these VNTR loci in forensic identification.
Short Tandem Repeats
Short tandem repeats (STRs) refer to repetitions of 2 to 5 bp.
Because STRs are so short, they are easy to amplify by
polymerase chain reaction (PCR), so RFLP analysis is not done
for STRs. DNA differing in one of two alleles is able to be
distinguished with great ease via STRs than with VNTRs.
Scientists add that STRs are plentiful—more than two thousand
are suitable for genetic mapping.9
In July 1993, Fregeau CJ et al came up with a new technique
for DNA typing. They analyzed eight different STRs by
amplifying with fluorescent tagged primers to directly analyze
the DNA typing on a fluorescent DNA fragment analyzer. In
their study a minimum of 100 picograms of DNA was required
for the procedure.10
According to Butler, also STRs have become more popular
for forensic typing because they are PCR-based and work with
low-quantity DNA samples. STRs are highly discriminating
between unrelated and even closely related individuals.2
METHODS OF DNA TYPING
Restriction Fragment Length Polymorphism (RFLP)
RFLP analysis is predicated on the existence of restriction
enzyme recognition sites that flank the polymorphic locus of
interest. The fidelity of restriction enzymes is such that DNA
restriction fragments are generated in a specific and reproducible
manner.
The basis of DNA profiling is the use of restriction fragment
length polymorphisms (RFLPs).
The three-dimensional structure of restriction enzymes
allows them to attach themselves to a double-stranded DNA
molecule and slide along the helix until they recognize a specific
sequence of base pairs which signals the enzyme to stop sliding.
The enzymes then digest (chemically separate) the DNA
molecule at that site—called a ‘restriction site’—acting like
molecular scissors, they cut DNA at a specific sequence of base
pairs. If a specific restriction site occurs in more than one
location on a DNA molecule, a restriction enzyme will make a
cut at each of those sites resulting in multiple fragments. The
length of each fragment will depend upon the location of
restriction sites contained within the DNA molecule.2
Electrophoresis then separates DNA fragments according
to their relative size. Over a period of time smaller fragments
will travel farther than larger ones. Fragments of the same size
stay together and migrate in single ‘bands’ of DNA.2
An RFLP probe is a labeled DNA sequence that hybridizes
with one or more fragments of the digested DNA sample after
they were separated by gel electrophoresis, thus revealing a
unique blotting pattern characteristic to a specific genotype at
a specific locus.
The RFLP probes are frequently used in genome mapping
and in variation analysis (genotyping, forensics, paternity tests,
hereditary disease diagnostics, etc.) (Fig. 2).
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Understanding the Basics of DNA Fingerprinting in Forensic Science

Polymerase Chain Reaction (Polymerase) Thus, samples used for extracting DNA have successfully
The PCR is an in vitro technique for the enzymatic amplification been reduced to a small piece of skin tissue, a single hair, or a
of small segments of DNA. PCR in its present form was invented drop of blood among other bodily fluids. If there is not a large
in 1985 and has been adapted more recently to forensic science. enough sample at a crime scene, PCR may be used to duplicate
The basic principle of PCR is based on the DNA replication. the DNA. Commonly, these DNA samples are used during crime
DNA polymerase enzyme uses the single stranded DNA (after scene investigations, paternity testing, military identification,
denaturation) as template for the synthesis of a new and animal poaching.
complementary strand and also requires a small section of Walsh DJ et al in their study have demonstrated that DNA
double-stranded DNA to initiate this synthesis. The starting banding patterns obtained from fresh saliva or various saliva-
point of DNA synthesis can be specified by supplying an stained materials, such as envelopes, buccal swabs, gags,
oligonucleotide primer that anneals (primer annealing ) to the cigarettes and forensic evidentiary samples containing mixed
template at a particular point. Both strands of DNA serve as saliva or semen stains were indistinguishable from patterns
templates for synthesis provided an oligonucleotide primer is obtained from blood or hair of same individual, concluding that
supplied for each strand. For PCR, the primers are chosen to saliva and saliva-stained material are good sources of DNA for
analysis.11
flank the regions of DNA that is to be amplified so that newly
synthesized strands of DNA, starting at each primer extend APPLICATIONS OF DNA FINGERPRINTING
beyond the position of the primers on the opposite strand (primer
extension). Thus, new primer sites are generated on each newly DNA fingerprintings have been used for all these following
scenarios:
synthesized DNA strand and cycle is repeated. Theoretically,
1. Accused and convicted felons can be set free because of
the DNA content is doubled with each complete cycle (Fig. 3).
DNA typing.
Thus, PCR is a powerful forensic diagnosis tool as it requires
2. Identifying of human remains.
only trace quantities (25 ng) of DNA and makes it realistic to
3. Determining relatedness of humans.
type the DNA from minute samples like even a single hair root
4. Studying relatedness among ancient people.
or a cigarette butt containing epithelial cells.5
PCR is most often used in STR analysis since the fragments
of interest are very short and easy to amplify. VNTR fragments
are longer and more difficult to amplify successfully.
PCR-based methods have rapidly overtaken RFLP
methods as:2
1. They are quicker
2. Need very little amount of DNA (0.1-1 ng)
3. Can be performed in degraded DNA
4. High power of discrimination
5. Automatable and capable of high volume sample processing.

Fig. 2: Process of RFLP analysis Fig. 3: Principle of PCR

Journal of Indian Academy of Oral Medicine and Radiology, October-December 2011;23(4):613-616 615
Vasavi Krishnamurthy et al
5. DNA testing of families.
6. Identifying organisms that cause disease.
7. Identifying birth parents (paternity testing).
8. Proving paternity.
9. Determining effectiveness of bone marrow transplants.
10. Proving relatedness of immigrants.
11. Confirming relatedness among animals.
In 1990, Haglund WD, Reay DT, Tepper SL published the
first report in the literature of a case using DNA fingerprinting
in a ‘parentage’ context to establish identity of unidentified
decomposed human remains. After routine methods failed to
establish positive identity of the victim, DNA typing techniques
using blood from the victim and putative parents of the victim
were used for establishing the identification.12
In 1992, Ginther C et al extracted mitochondrial DNA from
teeth stored from 3 months to 20 years. Blood from tooth donors
or their maternal relatives were taken for comparison. They
concluded that teeth provided an excellent source of DNA that
could be valuable for extending the time in which decomposed
human remains can be genetically identified.13
Yamamoto K in 1992 carried out study on DNA analysis
using tooth (dental pulp) as specimen along with age estimation
from tooth and inspection of dental and roentgenographic
records to provide rapid personal identification in major
disasters, traffic accidents, fires, etc.14
Once collected, the DNA should be preserved in a dry, cold
environment. While in the laboratory, cross contamination,
improper documentation, and nonstandardized procedures may
lead to greater loopholes and improper results.
Degradation and contamination are two major concerns in
handling potential DNA evidence. The breaking down of DNA
into smaller fragments by chemical or physical processes is
degradation, while contamination is the introduction of foreign
material to the sample. Potential causes of degradation are
overexposure to heat and humidity, leading to the growth of
bacteria, mold or mildew on a sample. Contamination can either
be the mixing of two DNA samples together, or when a person
inadvertently leaves a deposit in the evidence. Either
degradation or contamination can lead to false or un-
interpretable results.
In regard to this, teeth are resistant against extreme
circumstances, such as temperature, humidity and acidity, which
is an important advantage in DNA analysis. Furthermore,
abundance of DNA can be extracted from teeth.15
FUTURE OF DNA FINGERPRINTING
A number of automated or robotic instruments have been
developed recently that are capable of performing a variety of
tasks, ranging from rapid extraction robots to laboratory
automation workstation. One such workstation is Biomek 3000®
of Beckman Coulter Inc. has the ability to accurately and
616
reproducibly quantify the amount of amplifiable human DNA
from extracted DNA samples.16
In India, Indian biosciences offers a broad range of DNA
testing, paternity testing, genetic testing and DNA ancestry
services or DNA genealogy to provide indisputable answers to
emotional questions, professionally and confidentially.
The current techniques for analyzing DNA are somewhat
specialized, many future simplifications may promote ease of
usage. Better means to preserve samples of DNA in secure
facilities will hopefully make greater advances in the future.
The ability to cure genetically inclined diseases would be a
huge benefit to many suffering people for future generations.
The possibilities of DNA fingerprinting are endless. The use of
profiling has proven to be one of the greatest assets in the history
of science and a boon to forensic science.
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R (Ed). Forensic science review. Englewood Cliffs Prentice Hall,
1982;267-337.
4. Watson JD, Crick FHC. A structure of DNA. Nature 1953;171;
737-38.
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10. Fregeau CJ, Fourney RM. DNA typing with fluorescently
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12 Haglund WD, Reay DT, Tepper SL. Identification of
decomposed human remains by DNA profiling. J Forensic Sci
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13 Ginther C. Identifying individuals by sequencing mitochondrial
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14 Yamamoto K. Molecular biological studies on teeth inquests.
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