DNA FINGERPRINTING

By
DNA FINGERPRINTING1
Komal Kiran
Shreya Makala
WHAT IS DNA FINGERPRINTING

 DNA Fingerprinting is a technique employed

by forensic scientists to assist in the
identification of individuals on the basis of
their respective DNA profiles
 It uses repetative sequences that are highly
variable called Variable Number Tandom
Repeats [VNTR]
 It is also called DNA testing, DNA profiling ,
DNA typing or Genetic fingerprinting
 The chemical structure of everyone's DNA is
the same.
 The only difference between people (or any
animal) is the order of the base pairs.
 There are so many millions of base pairs in
eachperson's DNA that every person has a
different sequence
 Using these sequences, every person could
be identified by the sequence of their base pairs.
 Each person has a unique DNA fingerprint,
except monozygous (identical) twins.
 DNA fingerprint is the same for every cell,
tissue, and organ of a person. It cannot be
altered by any known treatment.
 Scientists use a small number of sequences of DNA
that are known to vary among individuals ,
and analyze those to get a certain probability of a matching
HOW IS IT PERFORMED

 Performing a southern blot

 Making a radioactive probe
 Creating a hybridization reaction
 VNTRs
SOUTHERN BLOT

The process involves :

 Isolating the DNA from the rest of the cellular
material in the nucleus.
 Cutting the DNA into several pieces of
different sizes
 Sorting the DNA pieces by size by
electrophoresis
 Denaturing the DNA to make it single
stranded
 Blotting the DNA to permanently attach
DNA to sheet
 It is analysed by the use of radioactive genetic
probe in hybridization with the DNA
 An X-Ray is taken of the blot after probe
bonds with the denatured DNA on the paper.
 Areas where the radioactive probe binds [red]
will show up on the film.
RADIOACTIVE BINDING
MAKING A RADIOACTIVE PROBE

 Obtain some DNA polymerase [pink]. Put the

DNA to be made radioactive (radiolabeled)
into a tube.
 Introduce nicks, or horizontal breaks along a
strand, into the DNA you want to radiolabel.
At the same time, add individual nucleotides
to the nicked DNA, one of which, *C [light
blue], is radioactive.
MAKING A RADIOACTIVE PROBE

 Add the DNA polymerase [pink] to the tube

with the nicked DNA and the individual
nucleotides. The DNA polymerase will
become immediately attracted to the nicks in
the DNA and attempt to repair the DNA,
starting from the 5' end and moving toward
the 3' end.
 The DNA polymerase [pink] begins repairing
the nicked DNA
 Whenever a G base is read in the lower strand, a
radioactive *C [light blue] base is placed in the new
strand. In this fashion, the nicked strand, as it is
repaired by the DNA polymerase, is made
radioactive by the inclusion of radioactive *C bases.
 The nicked DNA is then heated, splitting the two
strands of DNA apart. This creates single-stranded
radioactive and non-radioactive pieces. The
radioactive DNA, now called a probe [light blue], is
ready for use
CREATING A HYBRIDISATION REACTION

 Hybridization is the coming together, or

binding, of two genetic sequences. The binding
occurs because of the hydrogen bonds [pink]
between base pairs. Between a A base and a T
base, there are two hydrogen bonds; between a
C base and a G base, there are three hydrogen
bonds.
 When making use of hybridization in the
laboratory, DNA must first be denatured,
usually by using heat or chemicals
 Denaturing is a process by which the
hydrogen bonds of the original double-
stranded DNA are broken, leaving a single
strand of DNA whose bases are available for
hydrogen bonding
 Once the DNA has been denatured, a single-
stranded radioactive probe [light blue] can be
used to see if the denatured DNA contains a
sequence similar to that on the probe.
 The denatured DNA is put into a plastic bag
along with the probe and some saline liquid; the
bag is then shaken to allow sloshing. If the
probe finds a fit, it will bind to the DNA.
 The fit of the probe to the DNA does not have
to be exact. Sequences of varying homology
can stick to the DNA even if the fit is poor; the
poorer the fit, the fewer the hydrogen bonds
between the probe [light blue] and the
denatured DNA.
 The ability of low-homology probes to still
bind to DNA can be manipulated through
varying the temperature of the hybridization
reaction environment, or by varying the
amount of salt in the sloshing mixture.
VNTRs

 Every strand of DNA has pieces that contain

genetic information which informs an
organism's development (exons) and pieces
that, apparently, supply no relevant genetic
information at all (introns).
 Introns contain repeated sequences of base
pairs called VNTRs
 A given person's VNTRs come from the
genetic information donated by his or her
parents; he or she could have VNTRs
inherited from his or her mother or father, or
a combination, but never a VNTR either of his
or her parents do not have.
CASE STUDY

 Shown below are the VNTR patterns for Mrs.

Nguyen [blue], Mr. Nguyen [yellow], and their
four children: D1 (the Nguyens' biological
daughter), D2 (Mr. Nguyen's step-daughter,
child of Mrs. Nguyen and her former husband
[red]), S1 (the Nguyens' biological son), and
S2 (the Nguyens' adopted son, not
biologically related [his parents are light and
dark green]).
 Because VNTR patterns are inherited
genetically, a given person's VNTR pattern is
more or less unique. The more VNTR probes
used to analyze a person's VNTR pattern, the
more distinctive and individualized that
pattern, or DNA fingerprint, will be.
Use Of PCR

 PCR can be used in DNA fingerprinting as a

way to make numerous copies of isolated
DNA.  The process can selectively amplify a
single copy of a desired sequence
 Dna molecules are melted by raising the
temperature to 95 degrees Celcius.
 -Strands separate, temperature is lowered to
60 degrees Celcius.
 Each primer binds specifically to the 3 prime
end of the target sequence on the
appropriate strands of DNA.
 Primers direct taq polymerase to synthesize
complementary nucleotides.
 -Only DNA containing target sequence are
copied by taq polymerase. 
 At the end of cycle 1, both strands have been
copied to form 2 partially double-stranded
molecules
 These steps are repeated during the next
cycles.  After two cycles, four partially double-
stranded molecules are produced, all
containing the target sequence
 After many cycles, millions of copies of the
DNA can be produced. 
 -This is useful in DNA fingerprinting, as
researchers can do many tests on the same
DNA sample.  
PRACTICAL APPLICATIONS OF DNA
FINGERPRINTING

 Paternity and Maternity

Because a person inherits his or her VNTRs
from his or her parents, Parent-child VNTR
pattern analysis has been used to solve
standard father-identification cases as well as
more complicated cases of confirming legal
nationality and, in instances of adoption,
biological parenthood
 Criminal Identification and Forensics
DNA isolated from blood, hair, skin cells, or
other genetic evidence left at the scene of a
crime can be compared, through VNTR
patterns, with the DNA of a criminal suspect
to determine guilt or innocence. VNTR
patterns are also useful in establishing the
identity of a homicide victim, either from DNA
found as evidence or from the body itself.
 Personal Identification
The notion of using DNA fingerprints as a sort
of genetic bar code to identify individuals has
been discussed, but this is not likely to
happen anytime in the foreseeable future.
The technology required to isolate, keep on
file, and then analyze millions of very
specified VNTR patterns is both expensive
and impractical.
 Diagnosis of Inherited Disorders
 DNA fingerprinting is used to diagnose
inherited disorders in both prenatal and
newborn babies in hospitals around the
world. These disorders may include cystic
fibrosis, hemophilia, Huntington's disease,
familial Alzheimer's, sickle cell anemia,
thalassemia, and many others.
 Early detection of such disorders enables the
medical staff to prepare themselves and the
parents for proper treatment of the child. In
some programs, genetic counselors use DNA
fingerprint information to help prospective
parents understand the risk of having an
affected child
CASE STUDY

 In the example shown on the left, DNA

collected at the scene of a crime is
compared with DNA samples collected
from 4 possible suspects. The DNA has
been cut up into smaller pieces which are
separated on a gel. The fragments from
suspect 3 match those left at the scene of
the crime, betraying the guilty party.
THE END 