dr geeta j. gautam zoology department mmv, bhu

Dr Geeta J. Gautam

Zoology Department

MMV, BHU

• DNA fingerprinting is a test to identify and evaluate the

genetic information—called DNA (deoxyribonucleic acid)—in

a person's cells.

• It is called a "fingerprint" because it is very unlikely that any

two people would have exactly the same DNA information, in

the same way that it is very unlikely that any two people would

have exactly the same physical fingerprint.

• The test is used to determine whether a family relationship

exists between two people, to identify organisms causing a

disease, and to solve crimes.

• Only a small sample of cells is needed for DNA fingerprinting.

• A drop of blood or the root of a hair contains enough DNA for

testing. Semen, hair, or skin scrapings are often used in

criminal investigations.

DNA fingerprinting

DNA fingerprinting is done to:

1. Find out who a person's parents or siblings are. This test

also may be used to identify the parents of babies who

were switched at birth.

2. Solve crimes (forensic science). Blood, semen, skin, or

other tissue left at the scene of a crime can be analyzed to

help prove whether the suspect was or was not present at

the crime scene.

3. Identify a body. This is useful if the body is badly

decomposed or if only body parts are available, such as

following a natural disaster or a battle.

Why It Is Done

Blood sample from a vein

DNA that is used to establish identity is collected from a blood

sample.

Blood sample from a heel stick

If a DNA blood test is done on a baby, a heel stick will be

done instead of a blood draw from a vein.

For a heel stick blood sample, several drops of blood are

collected from the heel of the baby. The skin of the heel is

cleaned with alcohol and then pricked with a small, sterile

lancet. Several drops of blood are collected inside circles on a

specially prepared piece of paper.

Other methods

DNA can be collected from dried blood, skin, saliva, hair,

urine, and semen. Bone and teeth samples are used when a

body is badly decomposed.

Sample collection

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Restriction Enzymes

• A common use of restriction enzymes is to generate a "fingerprint" of a particular DNA molecule.

• DNA may be cut by some special naturally-occurring proteins called restriction enzymes.

• Each restriction enzyme only identifies and “cuts” at a very specific sequence (recognition site) in the DNA strand.

• Restriction enzymes typically recognise a symmetrical sequence of DNA, such as the site GAATTC.

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Example 1

• The top strand is the same as the bottom strand when you read backwards.

• When the enzyme EcoRI cuts the strand between G and A, it leaves overhanging chains:

• These are termed "sticky ends" because the base pairs formed between the two overhanging portions will glue the two pieces together, even though the backbone is cut.

Read to the left

Read to the right

Genetic fingerprinting

90% or more of DNA does not carry nucleotide triplets that code for proteins

The non-coding DNA is often called ‘junk DNA’ but this only means that its

functions have not yet been discovered

Some of the non-coding regions consist of repeated sequences of nucleotides

For example -C-A-T-G-C-A-T-G-C-A-T-G-C-A-T-G- *

The number of repeats in any one section of DNA varies from one

individual to the next

Since these sections do not code for proteins (and, therefore are not genes) there is

no observable difference in these individuals

Particular repeat sequences can be ‘cut out’ by restriction enzymes

For example

-CATCCACGACATGCATGCATGCATGCCACATCCA-

restriction enzyme cuts

here……………and…..….…..here

or

-CCACGACATGCATGCATGCATGCATGCATGCCACAT-

here…….…..…..………and…...….…………..here

Restriction enzymes

Gel electrophoresisThe different sized fragments are separated by a process called gel

electrophoresis

The separation takes place in a sheet of a firm but jelly-like substance

(a ‘gel’)

Samples of the DNA extracts are placed in shallow cavities (‘wells’)

cut into one end of the gel

A voltage is applied to opposite ends of the gel

DNA has a negative charge and moves slowly towards the positive end

The shorter fragments travel through the gel faster than the longer fragments

gelatinous sheet

well

solution

DNA extract

added

Gel electrophoresis

Voltage supply

negative electrode

DNA samples placed in

wells cut in gel

positive

electrode

thin slab of

gel

+DNA fragments

Move from negative

To positive

Gel electrophoresis

A sample with the

shorter DNA fragments

travels through the gel

faster than a sample

with the larger fragments

Outline of Electrophoresis

1. DNA fragments are loaded into “wells” in a gel. The gel floats in a buffer solution within a chamber between two electrodes.

2. When an electric current is passed through the chamber, negatively charged fragments move towards the positive terminal.

3. Shorter DNA fragments (smaller size) move faster than the longer ones (bigger size).

4. In a given time, DNA fragments are separated into bands according to their size.

“wells”

Source: Science Education Section, CDI, EDB.

Visualising DNA fragments in the gel

Appearance of separated fragments on gel

These bands will

contain the shorter

DNA fragments

These bands will

contain the longer

DNA fragments

starting positions Appearance of bands

© Prof. E. Wood© Prof. E.J.Wood


DNA analysis can be used for catching criminals, establishing

parentage, finding how closely organisms are related and many other

applications.

The pattern of bands in a gel electrophoresis is known as a

genetic fingerprint or a ‘genetic profile’

If a genetic fingerprint found in a sample of blood or other tissue

at the scene of a crime matches the genetic fingerprint of a suspect,

this can be used as evidence

A DNA sample can be obtained from the suspect using blood, cheek

epithelial cells taken from the mouth lining or even the cells clinging

to the root of a hair


….there is a chance of 1 in 10 that this

fragment occurs in many individuals…

Suppose that…………

…and.there is a chance of 1 in 20 that this





fragment occurs in many individuals, but…

Chances of a match

…the probability of all 4 bands matching in any person other thanthe suspect is

1 in 10 x 1 in 20 x 1 in 10 x 1 in 30

= 1 in 10 x 20 x 10 x 30 That is 1 in 60,000

When a larger number of bands is involved, the probability that the suspect is not guilty becomes one in many thousands*

Probability of a match

V S S1 S2 S3

V Victim

S Sample from crime scene

S1 Suspect 1

S2 Suspect 2

S3 Suspect 3

More than 20 fragments

from Suspect 1 match those

taken from the crime scene

DNA profiles

Evidence from genetic fingerprinting

Genetic fingerprinting is powerful evidence in criminal trials but…

• Many restriction fragments may be crowded into a single band

• There may be variability in the speed with which a fragment

travels through the gel

• There is a chance of contamination with ‘foreign’ DNA e.g. from

bacteria

• The jury may not understand the significance of genetic

fingerprinting and may be dependent on conflicting claims from

‘expert’ witnesses

• There may be arguments about the statistical significance of a

match between DNA profiles

Evidence from genetic fingerprinting

Even if there is agreement about a match between the

suspect’s DNA profile and forensic samples, it shows only

that the suspect was present at the scene of the crime and

does not prove that he or she committed the crime

DNA evidence should be considered as conclusive proof of

guilt only if there is other supporting evidence

In cases of paternity disputes, the genetic evidence can be

conclusive

Paternity can be decided on the basis of a single restriction site

Limitations of DNA evidence

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RFLP

• It was the first commercial technique of DNA analysis.

• It can be used in paternity cases or criminal cases to determine the source of a DNA sample.

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RFLP

• Steps

1. It uses restriction enzymes to cut DNA.

2. DNA fragments of different lengths are produced.

position of

restriction

fragment

part of DNA strand

mother father

Child will receive one copy of the restriction fragment from the

mother and one from the father. It could be any one of these

combinations

child

Paternity test 19

Starting position of sample

1 2 3 4

Genetic fingerprint of …

1 mother

2 child

3 possible father A

4 possible father B

There is a match between one of

the child’s restriction fragments

and one of the mother’s.

There is also a match between the

child’s other fragment and one from

possible father A.

Neither of the child’s restriction

fragments match those of possible

father B

Paternity test

20

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Activity DNA Fingerprint Analysis

Case 1

Mr. Chan’s family consists of mom, dad and four kids. The parents have one daughter and one son together, another daughter is from the mother’s previous marriage, and the other son is adopted. Here are the DNA analysis results:

1. Which child is adopted? Why?

2. Which child is from the mother’s previous marriage? Why?

3. Who are the own children of Mr and Mrs Chan?

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Activity 8.5

Answers:

• Child 4 is adopted.

• Child 2 is the child from the mother’s previous marriage.

• Child 1 and Child 3 are own children of Mr and Mrs Chan.

dr geeta j. gautam zoology department mmv, bhu

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