Copying DNA: Polymerase Chain Reaction

(PCR)

Created in 1983, Polymerase Chain Reaction (PCR) is a

process in which specialized polymerase enzymes are used
in the lab to artificially copy segments of DNA.
The process is done by a DNA Thermal Cycler and is highly
temperature dependent and involves extreme heating and
cooling. Each round of PCR doubles the amount of DNA.
After 30 rounds you have over a billion copies.
This process has revolutionized genetics because it allows
researchers to quickly amplify a small fragment of DNA in a
matter of a few hours.

DNA Profiling: STR Analysis

Introduction To Biochemcial Forensics

Paul G Hamm
What is DNA? Why analyze DNA?
DNA is a long polymer made up from monomers called
nucleotides. The four types of nucleotides are Adenine
(A), Guanine (G), Thymine (T), and Cytosine (C). These
bases make up the genetic alphabet and the order of
bases determines the structure of a protein.
The human genome contains around 3 billion base pairs
and close to 25,000 genes.
99.9% of human DNA is identical! This means the 0.1%
difference is what makes each human unique and this is
what forensic scientists can analyze in order to compare
DNA samples.

STR Analysis

PCR has dramatically changed DNA profiling because only a

small amount of DNA needs to be recovered from the crime
scene. (DNA profiles can be made from as little as 18 cells!)

Short Tandem Repeats (STR)

STRs are short repeating sequences in the DNA which are
3 to 7 bases long and are repeated in tandem at various
sites in the DNA. Each person has the same type of repeats
yet has a unique number of repeats at certain locations.
Example TH01 is the four letter A-A-T-G repeat.
At this site humans have from 5 to 11 of these repeats.
Example D7S820 is the four letter G-A-T-A repeat.
At this site humans have from 4 to 16 of these repeats.
In addition, humans have a maternal and paternal copy of
each gene. Thus humans have two sets of STRs at each
location. The exact number of STRs inherited at each
location makes a highly unique DNA profile.
Unlike other DNA markers, the entire strand of an STR is
relatively small (less than 450 bases) thus STRs are less
susceptible to degradation and can be recovered from
samples that have been subjected to decomposition.
**The images or photographs used herein may be copyright protected**

Once DNA is obtained from a stain or body fluid, the

DNA is first copied using PCR.
Various STR sites are studied by cutting the DNA into
fragments and analyzing them using gel electrophoresis.
An STR site with less repeats will travel farther through
the gel and a site with more repeats will travel a shorter
distance. Gel bands are easily visible and provide a quick
comparison.
If the bands match up at all of the comparison sites, that
increases the likelihood that the suspect DNA matches the
crime scene DNA.

Organizing DNA: Gel Electrophoresis

Gel electrophoresis is a process which allows researchers
to sort and analyze DNA fragments based on size.
DNA fragments are placed in an agarose gel and
subjected to an electric current.
Because DNA has a slightly negative charge, the DNA
fragments slowly travel through the gel towards the
positive electrode.

The smaller fragments move faster and thus farther while

the longer fragments move slower and thus they move
shorter distances.
A researcher can read the bands in the gel and analyze the
size of the fragments.

CODIS / Multiplexing
Using PCR and gel electrophoresis, forensic analysts can
compare multiple STR sites at the same time.
In the US, 13 STR sites are commonly used for DNA
profiling. The number of repeats at each site and their
relative frequency in the population has been studied and
the information is all gathered in the Combined DNA
Index System (CODIS).

Each STR is inherited independently, thus the probability

of having a particular combination of STR sites is the
product of the frequency of occurrence in the population.
While two people may have the same number of repeats
at one STR site, the probability that they have the same
combination of STRs at all 13 sites becomes very low.
Multiplying the frequencies of all 13 sites creates a DNA
profile that is highly unique and can be used with a high
degree of discrimination. Complete individualization can
often be achieved.
Comparing all 13 sites creates frequencies of occurrence
of 1 in 575 trillion for Caucasian Americans and
1 in 900 trillion for African Americans.