SUBJECT: SCIENCE OF CRIMINALISTICS

DATE: MARCH 24, 2018

REPORTER: MARIA KRISIA FAE R. DE ASIS
INSTRUCTOR: PROF. JUDITH M. BAGONA

DNA ANALYSIS
WHAT IS DNA?
Deoxyribonucleic acid, or DNA, is sometimes referred to as our genetic blueprint because it stores
the information necessary for passing down genetic attributes to future generations. Residing in
every nucleated cell of our bodies, DNA provides a “computer program” that determines our
physical features and many other attributes. The complete set of instructions for making an
organism is the entire DNA in a cell, referred to collectively as its genome.

DNA is a type of macromolecule known as a nucleic acid. It is organized into structures called
chromosomes and housed within the nucleus of our cells. DNA contains the genetic information
necessary for the production of other cell components and for the reproduction of life. Because the
DNA resides in the nucleus of the cell, it is often referred to as nuclear DNA. Some minor extranuclear
DNA, known as mitochondrial DNA, exists in the human mitochondria, the cellular powerhouses.

DNA is composed of a sugar and a phosphate, which form the backbone structure of the DNA
molecule, and four varying nucleobases: Adenine (A), Thymine (T), Cytosine (C), and Guanine (G). The
various combinations of these four letters, known as nucleotides, yield the diverse biological
differences among human beings and al living creatures.

The DNA, shaped like a double helix, are organized into chromosomes. Every cell contains forty-six
chromosomes. The father’s sperm contributes half of their chromosomes (23), while the mother’s
ova or egg contributes the other half (23). The human genome consists of 22 matched pairs of
autosomal chromosomes and two sex-determining chromosomes. Males are designated XY because
they contain a single copy of the X chromosome and single copy of the Y chromosome, while females
contain two copies of the X chromosome and are designated XX. Most human identity testing is
performed using markers on the autosomal chromosomes and gender determination is done with
markers on the sex chromosomes. The Y chromosome and mitochondrial DNA can also be used in
human identification applications.

A BRIEF HISTORY AND OVERVIEW OF FORENSIC DNA ANALYSIS

“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.

These DNA repeat regions became known as VNTRs, which stands for variable number of tandem
repeats. The technique used by Dr. Jeffreys to examine the VNTRs was called restriction fragment
length polymorphism (RFLP) because it involved the use of the restriction enzyme to cut the regions
of DNA surrounding the VNTRs. This RFLP method was first used to help solve the double homicide
case of Lynda Mann and Dawn Ashworth. Since that time, human identity testing using DNA typing
methods has been widespread.

FIRST USE OF FORENSIC DNA TESTING: CATCHING COLIN PITCHFORK

The first use of DNA testing in a forensic setting came in 1986. Two young girls, Lynda Mann and
Dawn Ashworth, were sexually assaulted and then brutally murdered in 1983 and 1986. Both murders
occurred near the village of Narborough in Leicestershire, England, with similar features, leading the
police to suspect that the same man had committed both crimes. Under public pressure, police
obtained a confession from a local man to killing one of the girls. His blood was compared to semen
recovered from the crime scenes. However, the man’s DNA did not match DNA evidence from either
crime. Thus, the first application of DNA technology was to demonstrate the innocence of someone
who might otherwise have been convicted.

A mass screening to collect blood for DNA testing from all adult men in three local villages was
conducted in a thorough search for the killer. More than 4000 men were tested without a match.
About a year later, a woman at a bar overheard someone bragging about how he had given a blood
sample for a friend named Colin Pitchfork. The police interviewed Mr. Pitchfork, collected a blood
sample from him, and found that his DNA profile matched semen from both murder scenes. He was
subsequently convicted and sentenced to life in prison.

The DNA typing methods used were Alec Jeffrey’s multi-locus RFLP probes, which he first described
in 1985. Since it was first used more than 20 years ago, DNA testing has progressed to become a
sensitive and effective tool to aid in bringing the guilty to justice and exonerating the innocent.

APPLICATIONS OF DNA TYPING

 Parentage Testing
 Disaster Victim Identification (DVI)
 Missing Persons Investigations
 Other uses: monitoring transplants, verifying cloning success, monitoring needle sharing
among drug users, detecting cancer tumors, mapping genetic diseases, tracing human
history, genetic genealogy

THE INNOCENCE PROJECT

Forensic DNA testing can play a role in protecting the innocent as well as implicating the guilty. In
recent years, the use of DNA evidence to free people from prison has been highly publicized and has
altered some perceptions of the criminal justice system. For example, capital punishment in Illinois
was put on hold in 2000 by the governor after learning of several inmates being exonerated by
postconviction DNA testing.

As of December 2008, a total of 225 people, including some “death row” inmates previously
incarcerated from crimes they did not commit, have been released from prison thanks to the power
of modern forensic DNA typing technologies. Many of these wrongfully convicted individuals were
found guilty prior to the development of DNA typing methods in the mid-1980s based on faulty
eyewitness accounts or circumstantial evidence. Fortunately for the more than 200 individuals
exonerated so far by postconviction DNA testing, some items from the crime scenes were preserved
in police evidence lockers that after many years could still be used for DNA testing. Results from
testing these old crime scene materials successfully excluded them as the perpetrator of the crimes
for which they were falsely convicted and imprisoned.

Defense attorneys Barry Scheck and Peter Neufeld launched the Innocence Project in 1992 at the
Benjamin N. Cardozo School of Law in New York City. This nonprofit legal clinic promotes cases
where evidence is available for postconviction DNA testing and can help demonstrate innocence.

The Innocence Project has grown to include an Innocence Network of more than 40 law schools and
other organizations around the United States and Australia. Law students and staff carefully evaluate
thousands of requests for DNA testing to prove prisoners’ innocence. In spite of careful screening,
when postconviction testing is conducted, DNA test results more often than not further implicate
the defendant. However, the fact that truly innocent people have been behind bards for a decade or
more has prompted legislation in a number of states and also at the federal level to fund
postconviction DNA testing.

BASIC PRINCIPLES OF FORENSIC DNA TYPING:

 The genome or the complete genetic composition of each individual is unique and is
inherited from an individual’s parents with one-half coming from the mother and one-half
from the father, with the exception of identical twins.
 A DNA profile by itself is fairly useless because it has no context. DNA analysis always
requires that a comparison be made between two samples: a questioned sample and a
known sample/

STEPS OF DNA TYPING USING SHORT TANDEM REPEAT (STR) MARKERS

Sample obtained from crime scene or paternity investigation

The different types of biological evidence collected at a crime scene can be used to associate or exclude an
individual from involvement with a crime. DNA evidence collection from a crime scene must be performed
carefully and a chain of custody established in order to produce DNA profiles that are meaningful and
legally accepted in court.
(See Table 1)

DNA Extraction
A biological sample obtained from a crime scene in the form of a blood or semen stain or a tissue (blood
or buccal swab) sample from a known individual contains a number of substances besides DNA. Therefore,
DNA molecules must be separated from other cellular material before they can be examined.
(See Table 2)
DNA extraction methods have been developed to separate proteins and other cellular materials from the
DNA molecules.
Extraction techniques: Organic Extraction, Chelex extraction, and solid-phase extraction or using FTA
paper (Flinder Technology Agreement)

DNA Quantitation
To ensure that DNA recovered from an extraction is human rather than from another source such as
bacteria, fungi, or animal, Quality Assurance Standards require human-specific DNA quantitation.
Typically, 0.5 to 2.0 ng of input human DNA is optimal for STR typing.

DNA Amplification (The Polymerase Chain Reaction or PCR)

DNA from crime scenes is often limited in both quantity and quality and obtaining a cleaner, more
concentrated sample is normally out of the question. First described in 1985 by Kary Mullis and members
of the Human Genetics group at the Cetus Corporation (now Roche Molecular Systems), PCR has
revolutionized molecular biology with the ability to make hundreds of millions of copies of a specific
sequence of DNA in a matter of only a few hours.
 Separation and Detection of PCR Products (STR Alleles)
A polymerase chain reaction (PCR) in which short tandem repeat (STR) alleles are amplified produces a
mixture of DNA molecules that present a challenging separation problem. A multiplex PCR can produce 20
or more different-sized DNA fragments representing different alleles that must be resolved from one
another. To distinguish the various molecules from one another, a separation step is required to pull the
different-sized fragments apart. The separation is typically performed by a process known as
Electrophoresis, which refers to electrical charges carried by the molecules.

Signals from fluorescent dyes attached to the DNA molecules can be used to detect and quantify the
relative amounts of DNA molecules being separated during electrophoresis.

Sample Genotype Determination

An STR Genotype is the allele present in a sample for a particular locus and is normally reported as the
number of repeats present in an allele. A full sample genotype or STR profile is produced by the
combination of all of the locus genotypes into a single series of numbers. This profile is what is being
entered into a case report or a DNA database for comparison purposes to other samples.

Comparison of Sample Genotype to other Sample Results

A DNA profile is made up of genotypes from individual genetic loci. The genotype at each locus results
from inheritance of paternal and maternal alleles.

Generally, the process of comparing two or more samples is limited to one of three possible outcomes
that are submitted in a case report:
(1) INCLUSION (MATCH) – peaks between the compared STR profiles have the same genotypes and
no unexplained differences exist between the samples.
(2) EXCLUSION (NONMATCH) – the genotype comparison shows profile differences that can only be
explained by the two samples originating from different sources.
(3) INCONCLUSIVE – the data does not support a conclusion as to whether the profiles match.

Comparison of DNA profile to Population Databases

When a match is observed between an evidence sample and a reference sample, then statistical methods
are typically invoked to provide information regarding the relevance of this match. When a DNA profile is
fairly common, then it is easier to imagine that the suspect might not be connected to the crime scene. On
the other hand, if the genotype is found to be extremely rare, then the evidence is stronger that the
suspect contributed the crime scene sample in question.

Generation of Case Report with Probability of Random Match

The end result of a forensic examination is a laboratory report, which represents a brief summary of a
work conducted by a forensic examiner or DNA analyst. The work represented in a laboratory report is
based on standard operating procedures that must be followed. Prior to release of lab report, data and
conclusions are vetted through an internal review process culminating with a second reviewer and/or the
DNA technical leader approving the work. This type of lab report is typically submitted to police
investigators to describe DNA typing results obtained from evidence and reference samples submitted.
Depending on the results, this report may also be used by a prosecuting attorney during court proceedings
to illustrate that a defendant’s DNA matches DNA evidence from a crime scene.
TABLE 1
 Some sources of biological materials used for PCR-based DNA typing
(Kuperus, et.al., 2003)

Automobile arm rest, baseball cap, bottle cap,

chocolate bar, cigarette lighter, cigarette paper,
credit card or ATM card, pistol magazine, coins,
doorbell, door pull, electrical cord, envelope,
DNA Source: HANDS gauze and tape, gloves, hammer, ignition
switch, keys, knife handle, hand-folded paper,
pen, plastic bag handles, remote car starter,
rope, screwdriver handle, seatbelt buckle, shoe
laces, steering wheel

Eaten apple, bile or vomit on sidewalk, bite

marks, bottle top, buccal swab, bitten food,
cigarette butt, envelope, glass rim, gum,
DNA Source: MOUTH and NOSE
inhaler, lipstick, nasal secretions, popsicle stick,
postage stamps, drinking straw, telephone
receiver, toothbrush, toothpick, utensils

Inside hat or cap, bullet, burned remains, buried

remains, contact lens, dandruff, umbilical cord,
DNA Source: GENERAL BODY eyeglasses, hair, hair comb, head-rest, jacket
hood, razor blade, tears, sweat, socks, toilet,
urine

TABLE 2
Typical DNA amounts that may be extracted from biological materials
(Lee and Ladd, 2001)
Type of Sample Amount of DNA
Liquid blood 20,000 – 40,000 ng/mL
Blood stain 250 – 500 ng/cm2
Liquid semen 150,000 – 300,000 ng/mL
Postcoital vaginal swab 10 – 3,000 ng/swab
Plucked hair (with root) 1 – 750 ng/root
Shed hair (with root) 1 – 10 ng/root
Liquid saliva 1,000 – 10,000 ng/mL
Oral swab 100 – 1,500 ng/swab
Urine 1 – 20 ng/mL
Bone 3 – 10 ng/mg
Tissue 50 – 500 ng/mg

Reference:

Butler, John M.
2010 Fundamentals of Forensic DNA Typing. National Institute of Standards and
Technology.