DNA FORENSICS
How does forensic identification work?
Any type of organism can be identified by examination of DNA sequences unique
to that species. To identify individuals, forensic scientists scan 13 DNA regions
that vary from person to person and use the data to create a DNA profile of that
individual (sometimes called a DNA fingerprint). There is an extremely small
chance that another person has the same DNA profile for a particular set of
regions.
Some Examples of DNA Uses for Forensic Identification
Identify potential suspects whose DNA may match evidence left at crime scenes
Exonerate persons wrongly accused of crimes
Identify crime and catastrophe victims
Establish paternity and other family relationships
Match organ donors with recipients in transplant programs
Is DNA effective in identifying persons?
Consider the scenario of a crime scene investigation
Assume that type O blood is found at the crime scene. Type O occurs in about
45% of Americans. If investigators type only for ABO, then finding that the
"suspect" in a crime is type O really doesn't reveal very much.
If, in addition to being type O, the suspect is a blond, and blond hair is found at
the crime scene, then you now have two bits of evidence to suggest who really
did it. However, there are a lot of Type O blonds out there.
If you find that the crime scene has footprints from a pair of Nike Air Jordans (with
a distinctive tread design) and the suspect, in addition to being type O and blond,
is also wearing Air Jordans with the same tread design, then you are much closer
to linking the suspect with the crime scene.
In this way, by accumulating bits of linking evidence in a chain, where each bit by
itself isn't very strong but the set of all of them together is very strong, you can
argue that your suspect really is the right person.
How is DNA typing done?
•About 3 million bases differs from one person to the next.
•Scientists can use these variable regions to generate a DNA profile of an
individual, using samples from blood, bone, hair, and other body tissues and
products.
In criminal cases, this generally involves obtaining samples from crime-scene
evidence and a suspect, extracting the DNA, and analyzing it for the presence of a
set of specific DNA regions (markers).
•Scientists find the markers in a DNA sample by designing small pieces of DNA
(probes) that will each seek out and bind to a complementary DNA sequence in
the sample. A series of probes bound to a DNA sample creates a distinctive
pattern for an individual.
•Forensic scientists compare these DNA profiles to determine whether the
suspect's sample matches the evidence sample. A marker by itself usually is not
unique to an individual; if, however, two DNA samples are alike at four or five
regions, odds are great that the samples are from the same person.
•If the sample profiles don't match, the person did not contribute the DNA at the
crime scene.
•The more probes used in DNA analysis, the greater the odds for a unique pattern
and against a coincidental match, but each additional probe adds greatly to the
time and expense of testing.
•A minimum of six probes are recommended.
DNA chip technology (in which thousands of short DNA sequences are embedded
in a tiny chip) will enable much more rapid analysis using many more probes, and
raising the odds against coincidental matches.
Some of the DNA technologies used in forensic
investigations
Restriction Fragment Length Polymorphism (RFLP)
RFLP is a technique for analyzing the variable lengths of DNA fragments that result from
digesting a DNA sample with a special kind of enzyme. This enzyme, a restriction
endonuclease, cuts DNA at a specific sequence pattern know as a restriction
endonuclease recognition site. The presence or absence of certain recognition sites in a
DNA sample generates variable lengths of DNA fragments, which are separated using
gel electrophoresis. They are then hybridized with DNA probes that bind to a
complementary DNA sequence in the sample.
RFLP is one of the original applications of DNA analysis to forensic investigation. With
the development of newer, more efficient DNA-analysis techniques, RFLP is not used as
much as it once was because it requires relatively large amounts of DNA. In addition,
samples degraded by environmental factors, such as dirt or mold, do not work well with
RFLP.
PCR Analysis
PCR (polymerase chain reaction) is used to make millions of exact copies of DNA from a
biological sample. DNA amplification with PCR allows DNA analysis on biological
samples as small as a few skin cells. The ability of PCR to amplify such tiny
quantities of DNA enables even highly degraded samples to be analyzed.
Great care, however, must be taken to prevent contamination with other biological
materials during the identifying, collecting, and preserving of a sample.
STR Analysis
Short tandem repeat (STR) technology is used to evaluate specific regions (loci) within
nuclear DNA. Variability in STR regions can be used to distinguish one DNA profile from
another. The Federal Bureau of Investigation (FBI) uses a standard set of 13
specific STR regions for CODIS. CODIS is a software program that operates local,
state, and national databases of DNA profiles from convicted offenders, unsolved
crime scene evidence, and missing persons.
The odds that two individuals will have the same 13-loci DNA profile is about one
in one billion.
STRs are currently the most popular type of DNA fingerprint, since the whole
PCR process takes only a few hours, compared to RFLP/VNTR probe
hybridization and film exposure which can take several days. STRs can use much
smaller samples of DNA than RFLPs/VNTRs, and can even use partially degraded
DNA to create a fingerprint.
Thus, the integrity and quality of the DNA sample is not as great a factor with
STRs than with the traditional methods of DNA fingerprinting.
The current standard forensic protocol analyses 13 core STR loci which have
been carefully chosen for their uniqueness. The only disadvantage of the STR
approach is it is sensitive to contaminating DNA, so usually the STR approach is
used first, followed by a VNTR analysis if contamination is suspected, and enough
DNA is available.
Mitochondrial DNA Analysis
Mitochondrial DNA analysis (mtDNA) can be used to examine the DNA from
samples that cannot be analyzed by RFLP or STR. Nuclear DNA must be extracted
from samples for use in RFLP, PCR, and STR.
While older biological samples that lack nucleated cellular material, such as hair,
bones, and teeth, cannot be analyzed with STR and RFLP, they can be analyzed
with mtDNA. In the investigation of cases that have gone unsolved for many
years, mtDNA is extremely valuable.
All mothers have the same mitochondrial DNA as their daughters. This is because
the mitochondria of each new embryo comes from the mother's egg cell.
The father's sperm contributes only nuclear DNA.
Comparing the mtDNA profile of unidentified remains with the profile of a
potential maternal relative can be an important technique in missing person
investigations.
Y-Chromosome Analysis
The Y chromosome is passed directly from father to son, so the analysis of
genetic markers on the Y chromosome is especially useful for tracing
relationships among males or for analyzing biological evidence involving multiple
male contributors.
VNTRs - σημαντικοί γενετικοί μάρτυρες για εγκληματολογικές αναλύσεις
•Το μέγεθος του ενισχυμένου τμήματος ή αλληλομόρφου εξαρτάται από τον
αριθμό και το μέγεθος των επαναλαμβανόμενων αλληλουχιών που περιέχονται σε
αυτό.
•Με κατάλληλους VNTR γενετικούς τόπους και υψηλής διακριτικής ικανότητας
ηλεκτροφορητικά συστήματα, η ενίσχυση συγκεκριμένων VNTR αλληλουχιών με
PCR μπορεί να φανεί χρήσιμη για αναλύσεις ταυτοποίησης.
Legend:
(A) The DNA sequences that create the variability used in this analysis contain
runs of short, repeated sequences, such as GTGTGT . . . , which are found in
various positions (loci) in the human genome. The number of repeats in each
run is highly variable in the population, ranging from 4 to 40 in different
individuals. A run of repeated nucleotides of this type is commonly referred to
as a VNTR (variable number of tandem repeat) sequence. Because of the
variability in these sequences, each individual will usually inherit a different
variant of each VNTR locus from their mother and from their father; two
unrelated individuals will therefore not usually contain the same pair of
sequences.
A PCR reaction using primers that bracket the locus produces a pair of bands of
amplified DNA from each individual, one band representing the maternal
variant and the other representing the paternal variant. The length of the
amplified DNA, and thus its position after electrophoresis, will depend on the
exact number of repeats at the locus.
(B) The DNA bands obtained from a set of four different PCR reactions, each of
which amplifies the DNA from a different VNTR locus. In the schematic
example shown here, the same three VNTR loci are analyzed from three
suspects (individuals A, B, and C), giving six bands for each person. You can
see that although some individuals have several bands in common, the overall
pattern is quite distinctive for each.
The band pattern can serve as a “fingerprint” to identify an individual nearly
uniquely.
The fourth lane (F) contains the products of the same PCR reactions carried out
on a forensic sample. The starting material for such a PCR reaction can be a
single hair that was left at the scene of the crime. From the example, individuals A
and C can be eliminated from enquiries, while B remains a clear suspect.
One of the first accepted uses of DNA fingerprinting was in the investigation of
sexual assault and rape cases. Detectives only had to match the DNA of the semen
found
at the scene of the crime with the DNA of any potential suspect to determine who was
guilty of committed the crime. A DNA sample from the rapist could be obtained from a
simple vaginal swab from the victim or any other semen that was released in the area
during the assault.
The figure below shows how a DNA fingerprint can help determine
who is guilty of a sexual assault.
As seen from figure below , suspect B (lane 4) is guilty of rape because his DNA
fragments
match that of the semen found on the victim’s clothes (lane 3) and also in the vagina
(lane 6). Suspect A (lane 2) is clearly not the rapist because his DNA fragments do not
match the semen found on the victim’s clothes or the semen from the vaginal swab.
DNA fingerprinting is very useful in such an application because it provides the police
with an exact match of who left evidence at the crimescene.
In paternity tests potential fathers of the child have their
DNA analyzed with the child and mother’s DNA in order to see which of the
potential fathers has the most DNA in common with the child in question. Figure 6
shows an example of a RFLP used to determine which potential father (F1 and F2)
is the real father of the child (C). As you can see in the figure below, the second
father tested (F2) seems to have more DNA in common with the child than that of
the first father tested (F1).
Figure 6.
A
paternity
test using
the RFLP
technique
DNA can be obtained from traces of biological fluids or tissues at a crime scene.
The most traditional sources of forensic DNA come from saliva, seminal fluid,
blood, and hair. DNA from saliva can be extracted from items such as cigarette
butts, ski masks, envelopes, and stamps. Seminal fluids are usually found from
oral, rectal, or vaginal swabs, and also on clothing. Blood stains and hair that are
visible at a crime scene usually contain DNA that can be analyzed at the forensic
laboratory
Contamination is one of the greatest risks that the evidence must be guarded
from.If your sample of evidence is found to be contaminated, it can be thrown out
as evidence in the courtroom. Contamination can occur at the crime scene, during
packaging, in transit to the laboratory, and also during analysis. With a risk of
possible contamination present in all these steps of the forensic process, proper
precautions must be used to prevent ruining the DNA sample.
At the crime scene many factors must be considered in trying to prevent
contamination. The first factor is Nature. For example, if it rained at the crime
scene,a blood stain found could be diluted which would be almost impossible to
analyze. Also if it was windy that day then vital pieces of DNA could have been
blown away from the crime scene.
Another factor at the crime scene is properly securing the area so that people do
not taint the evidence. Until a crime scene is secured many individuals not related
to the event may have left DNA around key evidence which may
be mistaken for a possible suspect. Equipment is another factor which must be
regulated to reduce the risk of evidence contamination.
Clothing, notepads, photography equipment, and crime scene kits must be
properly decontaminated once leaving a crime scene or they may contaminate
evidence at another crime scene.
Disposable personal protective equipment (PPE) should be worn including: a
mask, jumpsuit, gloves, booties and head cover.
By keeping these tips in mind, contamination at a crime scene should be at a
minimum.
During packaging and transport of the DNA evidence, there are also a few factors
to keep in mind in order to preserve the sample in its original state. When
packaging evidence, each sample should be sealed individually to prevent crosscontamination.
Biological fluids should be dried in order to avoid contamination by bacteria .
Evidence should be packaged in a paper container, which is then put into an open
plastic container to reduce the risk of evidence leaking out while in transit.
When the biological sample is in transit certain precautions must be taken. DNA is
sensitive to temperature which means that while the sample is in transit, the
package must be out of direct sunlight or any other extreme circumstance.
The samples acquired from the crime scene should be kept away from high
temperatures. Room temperature is acceptable, refrigeration is desirable, and
freezing is preferred when 20 it comes to storing a sample.
DNA at an Aged Crime Scene
At an aged crime scene, forensic scientists attempt to retrieve DNA evidence that
is the least contaminated and decomposed. For example, if a deceased individual
is at an aged crime scene and is showing signs of decomposition, then blood
work would not be a practical route of obtaining a valid DNA sample .
If hair is present on the deceased body, the sample must be pulled from the tissue
because the tissue attached to the hair root is necessary for DNA analysis.
Bones and teeth would also serve as a suitable sample of DNA evidence, since
they take very long to degrade.
We now know that tiny particles of biological fluids, most notably blood, can cling
to most surfaces at the crime scene for many years without anyone ever realizing
they are there . There is now a chemical that allows you to see these microscopic
particles of blood and find the clues, that to the naked eye,did not seem to exist.
The chemical is called luminol and it reveals these particles with a
light-producing chemical reaction between hemoglobin and several chemicals.
Light is produced because the reactants contain more energy than the products
of the reaction, which release the extra energy in the form of light photons. This is
the process, which is called chemiluminescence. After the chemical
is sprayed onto the object in question, a blue glow will emit in any areas where
there are
tiny traces of blood, an example of which is shown in figure 4.
A simulation of luminol at work. The chemical binds to the blood and emits a blue
glow. Once the luminol reacts with the small traces of blood left behind, forensic
investigators will takes pictures or videotape the crime scene to record any
potential patterns . It should be noted that once blood reacts with luminol, that
sample is no longer able to provide DNA for analysis. Perhaps in the future a new
method of visualization will be devised that maintains DNA structure.
CONCLUSIONS
DNA fingerprinting is the most sophisticated way to identify living organisms.
DNA cannot easily be altered once it is left at a crimescene or deposited with a
mummy, which makes it a strong forensic tool.
RFLPs and VNTRs are the traditional methods of fingerprinting DNA, which uses a
relatively large sample that uses the method of probe hybridization to detect
polymorphisms in the DNA.
STRs are the most current form of DNA fingerprinting, which is PCR based and
uses a very small sample of DNA. DNA fingerprinting has many applications that
range from criminal rape cases, paternity tests, molecular archeology etc.
DNA forensics is one of the greatest tools in piecing together a crime scene. Over
the past ten years there have been many advances in the methods of collecting
and preserving these DNA samples to help facilitate the acceptance of this
evidence in the court room. By avoiding contamination and properly storing it to
prevent degradation,forensic science has made a monumental step in allowing
DNA samples as valid evidence in United States courtrooms. DNA evidence is
now one of the most powerful tools used in determining who is responsible for a
crime. With criminals altering their fingerprints and other physical characteristics,
DNA evidence is one of the only true methods to correctly identify an individual.
Now with the help of chemicals such as luminol, crime scenes that at first
analysis seem to have no physical evidence are further examined on the particle
level which makes it almost impossible to leave a crime without a trace.
DNA Forensics Databases
National DNA Databank: CODIS
The COmbined DNA Index System, CODIS, blends computer and DNA technologies
into a tool for fighting violent crime. The current version of CODIS uses two indexes to
generate investigative leads in crimes where biological evidence is recovered from the
crime scene. The Forensic index contains DNA profiles developed from crime scene
evidence. All DNA profiles stored in CODIS are generated using STR (short tandem
repeat) analysis.
CODIS utilizes computer software to automatically search its two indexes for matching
DNA profiles. Law enforcement agencies at federal, state, and local levels take DNA
from biological evidence (e.g., blood and saliva) gathered in crimes that have no suspect
and compare it to the DNA in the profiles stored in the CODIS systems. If a match is
made between a sample and a stored profile, CODIS can identify the perpetrator.
All 50 states have laws requiring that DNA profiles of certain offenders be sent to
CODIS. As of January 2003, the database contained more than a million DNA
profiles in its Convicted Offender Index and about 48,000 DNA profiles collected
from crime scenes but which have not been connected to a particular offender.
.
In March 2003 USA goverment proposed $1 billion in funding over 5 years to
reduce the DNA testing backlog, build crime lab capacity, stimulate research and
development, support training, protect the innocent, and identify missing
persons.
More on CODIS
CODIS: Combined DNA Index System - Information from the FBI.
The FBI Laboratory's Combined DNA Index System Program - Enter regional information to learn more
about CODIS in your area of the world. From Promega Corporation, a major supplier of reagents and other
materials to support molecular biology research.
National Commission on the Future of DNA Evidence
Postconviction DNA Testing: Recommendations for Handling Requests - Report from the National
Commission on the Future of DNA Evidence
Another application of DNA fingerprinting is a more recent method in molecular
archeology. This method of archeology uses DNA to determine a species of an
archeological discovery or to trace blood lines of animal or human remains. DNA
may be extracted from biological remains, hair, teeth, body tissues, or even
fossils.
The best climates to preserve DNA are very cold temperatures and arid climates.
Some examples of specimens from these types of climates are the “Tyrolean IceMan”, who was found in the Alps, and the mummies of Egypt found in the dry
desert.
The ice man was found to be around 5300 years old, and DNA was extracted from
the remains of his gut which found small traces of food that he ate (Ice Man,
2005).
This was one of the most historic archeological discoveries in the last century.
DNA fingerprinting is an important tool for archeologists to piece together
information that links the past to us today.