A brief account of DNA fingerprinting, its applications and limitations

1. DNA FINGERPRINTING
Dr. Saji Mariam George
Associate Professor
Assumption College Autonomous
Changanacherry

2. DNA FINGER PRINTING
(DNA PROFILING, DNA TYPING, MOLECULAR FINGERPRINTING, GENETIC
FINGERPRINTING)
• One of the applications
of Biotechnology - in
Forensics.
• Developed by
Alec Jeffreys (1985).

3. • DNA fingerprints are specific banding patterns
on Southern blots of genomic DNA cleaved with
a specific restriction enzyme and hybridized to
appropriate DNA probes – Autoradiogram is
taken to get a DNA print pattern of the
individuals.
• From the DNA print pattern, the individuals are
distinguished on the basis of bands in their DNA
prints .
• An individual can be identified at molecular
level.

4. • Human genome - 3billion nucleotides.
• No two individuals have the same nucleotide
sequences.
• The uniform nature of DNA in a single
individual and the genetic variability
between individuals make DNA finger
printing possible.

5. • DNA finger printing relies not on full genome
sequencing , but on identifying differences in some
specific regions in DNA called repetitive sequences ( a
small stretch of DNA is repeated many times) –
satellite DNA or microsatellites or minisatellites –
highly variable : Variable Number Tandem
Repeats(VNTRs).
• Copy numbers varies from chromosome to
chromosome in an individual.
• Each organism has a unique pattern of VNTRs - can
provide valuable evidence in cases of uncertain
identity.

6. Variable Number Tandem Repeats(VNTRs).
Image:https://slideplayer.com/slide/14307419

7. • VNTRs are very similar between closely related
humans, but dissimilar between unrelated
individuals.
• Do not code for any proteins – form a large
portion of human genome.
• Sequences show high degree of polymorphism
due to mutations and form the basis of DNA
fingerprinting- Comparing a number of VNTRs in
a given area, one can identify a person easily.
(DNA polymorphism – an inheritable mutation
in a population at high frequency).

8. Steps in DNA Fingerprinting
1. Isolation of cell DNA.- DNA is isolated from
the source materials – a drop of blood,
semen, teeth, bones, hair follicle, tissues ,
suspected individuals etc.
1. Digestion of DNA by restriction
endonuclease – DNA is cut with a restriction
enzyme that cut on either side of a
minisatellite(VNTRs) .

9. 3. PCR amplification – Since samples provide a
small amount of DNA, it is amplified by PCR
(Polymerase Chain Reaction).
4. Separation of DNA fragments by Gel
electrophoresis
Each DNA restriction digest of the resource
persons and the source material is poured
into the wells of an electrophoresis gel and
electrophoresis is carried out.

10. • The minisatellites separate according to
their lengths. The gel is chemically treated
(alkali treatment – 0.5 M NaOH ) or heated to
denature the DNA to form single strands.
ssDNA (single stranded DNA) is capable of
binding to single stranded probes that can be
used to detect unique DNA sequences.

11. 5.Southern blotting and baking – the separated
DNA fragments are transferred to a nylon
membrane or a nitrocellulose filter paper by
placing it over a gel. The nitrocellulose filter
having DNA fragments is dried in between
dry filter papers at high temperature
(Baking).

12. 6. Selection of DNA probe
• Single stranded fragments of DNA containing
the complementary code for a specific
sequence of bases.
• The targeted area on the DNA sample is
called a locus. A single locus probe targets a
sequence that appears in only one position
on the genome. A multi locus probe attaches
to sequences in many places in the genome.

13. • DNA probes developed by Alec Jeffreys are
used for DNA finger printing.
• In India, BKm probe – developed from mini
satellite DNA of Banded Krait by Dr Lalgi
Singh , Centre for Cellular and Molecular
Biology, Hyderabad.

14. 7. Hybridization with labelled VNTR probe
(Filter Hybridization)
• Binding of the DNA fragment with the
complementary probe - Nitrocellulose filter
is placed in hybridization solution containing
the radioactively labelled probe – The probe
DNA binds with appropriate minisatellites
and form duplex DNAs.

15. 8. Detection of hybridized DNA fragments by
Autoradiography
• After hybridization, the filter is washed with a wash
solution to remove unbound probes.
• An X-ray film is placed over the filter for about 3
hours – The radioactivity of the probes makes dark
spots on the X-ray film – an irregular ladder of dark
spots develops on the X-ray film for each DNA sample
– Each ladder represents a DNA print of an individual.
All DNA prints on the X-ray film together constitute a
DNA finger print pattern.

16. 9. Analysis of DNA print pattern.
• Compare two or more autoradiographs to see
the band match.
• Based on the degree of band match, we can
arrive at a conclusion regarding the
identification of an individual.

17. Applications of DNA Fingerprinting
1. Forensic applications
• A powerful forensic tool - DNA finger
prints(VNTR prints) were first used as evidence
in criminal case in 1988.
• DNA from any part of the body - hair follicle,
semen, a drop of blood etc. from the crime
scene is enough to establish innocence or guilt.

18. • Can be used to identify a criminal – a
murderer, rapist etc. whose DNA may match
evidence left at crime scenes.
• It can also be used to identify a victim in
cases where the body may be disfigured and
teeth or other identifying features may be
destroyed.

19. • An analysis of the DNA
print pattern shows that
the bands of the
specimen and the suspect
No. 1 match. Hence he is
the criminal. The suspects
No. 2 & 3 are not guilty .
Image:https://prezi.com/

20. 2. To settle disputed Parentage
(Paternity or maternity tests).
• DNA samples are obtained from the cells of
the child, the mother and possible fathers
and DNA finger prints were prepared.
• When the DNA fingerprints were compared,
all the bands in the child’s DNA print should
be present in the combined DNA prints of
the parents.

21. • For each pair of homologous chromosomes ,
the child will have received one from each
parent - Thus approximately, half of the
bands in the child’s DNA print will result
from DNA sequences inherited from the
mother and the other half from the DNA
sequences inherited from the father.
• The accuracy can be increased by using
more probes – more DNA polymorphisms
can be surveyed and a large proportion of
the child’s DNA and parents’ DNA can be
compared.

22. • A critical analysis of the
band match revealed that
the bands of the child and
the alleged father on the
fingerprint pattern on the
right side match – hence
he is the biological father.
(Paternity inclusion).

23. 3.DNA fingerprints can be
used to determine
genetic family
relationships
( sibling relationships or
other kinships)
Example
Daughter 1 and son 1 share
RFLPs (band match ) with
both Mom & Dad (Coloured
blue & yellow
respectively).(RFLP-
Restriction Fragment Length
Polymorphism)
Daughter 2 has RFLP of Mom ,
but not the Dad, D2 - is the
child from mothers previous
marriage .
Son 2 does not have RFLPs from
either parent – Son 2 is
adopted.
Image: http://mol-biol4masters.masters.grkraj.org/

24. 4. Personal Identification
 DNA fingerprints can be used to generate
DNA profile of an individual.
 Can be used as a type of genetic barcode to
identify individuals.

25. Limitations of DNA Fingerprinting
1. Requires further standardization and
quality control to be accepted as a tool.
2. There are only a few labs around the
world that can give accurate results.
3. There is 1 in 50 billion chance of two
DNA sequences being similar.

26. 4. The probability of human error in producing
prints and the methods for calculating the
probability that two individuals have identical
fingerprints – researchers must have reliable
information about the frequency of the DNA
polymorphism in the population in
question – if inbreeding is common in the
population, the probability of identical
fingerprints will increase.

27. THANK YOU