CERTIFICATE

This is certified that Udit, student of XII Medical has completed the

project “DNA Fingerprinting”. The work done in the project is the result of

candidate’s own efforts. The project is considered as a part of fulfillment of

Biology-Practical Examination of 10+2 class.

Mrs Shalini Chaudhary

Lecturer in Biology
 ACKNOWLEDGEMENT

I have great pleasure in recording my profound gratitude to

Mrs. Shalini Chaudhary, for her invaluable guidance, constant

encouragement and immense help given at each step for persuing this work

which reveals his vast knowledge in the field of Biology.

Udit
Class – XII, Medical
 CONTENTS

 Meaning of DNA Fingerprinting

 Historical Aspect

 Principle of DNA Fingerprinting

 Technique for DNA Fingerprinting

 Applications of DNA Fingerprinting

 Problems
 DNA FINGERPRINTING

What is DNA 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 each persons DNA that every

person has a different sequence.

Using these sequences, every person could be identified solely by the

sequence of their base pairs. However, because there are so many millions of

base pairs, the task would be very time-consuming. Instead, scientists are

able to use a shorter method, because of repeating patterns in DNA.

These patterns do not, however, give an individual ‘fingerprint,’ but

they are able to determine whether two DNA samples are from the same

person, related people, or non-related people. Scientists use a small number

of sequences of DNA that are known to vary among individuals a great deal,

and analyze those to get a certain probability of a match.

DNA fingerprinting (=DNA typing) is a technique to identify a person

on the basis of his/her DNA specificity. Every individual organism is

unique. Each person has a unique DNA fingerprint. Unlike a conventional

fingerprint. Unlike a conventional fingerprint that occurs only on the

fingerprints and can be altered by surgery, a DNA fingerprint is the same for

every cell, tissue and organ of a person. It cannot be changed by any known

treatment. The ideal way to distinguish other people whould be his or her

entire genomic DNA sequence.

DNA fingerprints is a very quick way to compare the DNA sequences

of any two individuals.

If one aims to find out genetic differences b/w 2 individuals or among

individuals of a population sequencing the DNA every time would be a

daunting and expensive task.

 HISTORICAL ASPECT

The Study of finger, palm and role prints is called dermatoglyphics. It

has been a subject of human interest since times when man used to went for

his food with the help of animal’s foot prints. Science of fingerprinting was

first used by Sir William Herschel as a method of identification in 1858. In

India the science of fingerprints was discovered by chance during a murder

investigator in Jalpaiguni in 1897.

Sir Alec Jeffreys (1984) invented the DNA, fingerprinting technique

at Leicester University, United Kingdom Dr. V.K. Kashyap and Dr. Lalji

Singh started the fingerprinting technology in India.

 PRINCIPLE OF DNA FINGERPRINTING

DNA fingerprinting involves identifying differences in some specific

regions in DNA sequence called as repetitive DNA, because in these

sequences, a small stretch of DNA is repeated many times. These repetitive

DNA are separated from bulk genomic DNA as different peaks during

density gradient centrifugation. The bulk DNA forms a major peak and the

other small peaks are referred to as satellite DNA. Depending on base

composition (A: T rich or G:C rich), length of segment, and number of

repetitive units, the satellite DNA is classified into many categories, such as

micro-satellites, mini-satellites etc. These sequences normally do not code

for any proteins, but they form a large portion of human genome. These

sequence show high degree of polymorphism and form the basis of DNA

fingerprinting. Since DNA from every tissue (such as blood, hair-follicle,

skin, bone, saliva, sperm etc.), from an individual show the same degree of

polymorphism, they become very useful identification tool in forensic

applications. Further, as the polymorphisms are inheritable from parents to

children. DNA fingerprinting is the basis of paternity testing, in case of

disputes.
 As polymorphism in DNA sequence is the basis of genetic mapping of

human genome as well as of DNA fingerprinting, it is essential that we

understand what DNA polymorphism means in simple terms. Polymorphism

(variation at genetic level) arises due to mutations. (Recall different kind of

mutations and their effects. New mutations may arise in an individual either

in somatic cells or in the germ cells (cells that generate gametes in sexually

reproducing organisms). If a germ cell mutation does not seriously impair

individual’s Ability to have offspring who can transmit the mutation, it can

spread to the other members of population (through sexual reproduction).

Allelic sequence variation has traditionally been described as a DNA

polymorphism if more than one variant (allele) at a locus occurs in human

population with a frequency greater than 0.01. In simple terms, if an

inheritable mutation is observed in a population at high frequency, it is

referred to as DNA polymorphism. The probability of such variation to be

observed in non-coding DNA sequence would be higher as mutations in

these sequences may not have any immediate effect/impact in an

individual’s after generation, and form one of the basis of

variability/polymorphism. There is a variety of different types of

polymorphisms ranging from single nucleotide, such polymorphisms play

very important role, and you will study these in details at higher classes.
 The technique of DNA fingerprinting was initially developed by Alec

Jeffreys. He used a satellite DNA as probe that shows very high degree of

polymorphism. It was called as Variable Number of Tandem Repeats

(VNTR).

The VNTR belongs to a class of satellite DNA referred to as mini-

satellite. A small DNA sequence is arranged tandemly in many copy

numbers. The copy number varies from chromosome to chromosome in an

individual. The numbers of repeat show very high degree of polymorphism.

As a result the size of VNTR varies in size from 0.1 to 20kb. Consequently,

after hybridization with VNTR probe, the autoradogram gives many bands

of differing sizes. These bands give a characteristic pattern for an individual

DNA. It differs from individual to individual in a population except in the

case of monozygotic (identical) twins. The sensitivity of the technique has

been increased by use of polymerase chain reaction.

 TECHNIQUE FOR DNA-FINGERPRINTING

1) The DNA is extracted from the nuclei of white blood cells or of

spermatozoa or of the hair follicle cells that cling to the roots of hairs

that have fallen, or been pulled out.

2) The DNA molecules are first digested with the help of enzyme

restriction endonuclease that cuts them into fragments. The fragments

of DNA also contain the VNTRs.

3) The fragments are separated according to size by gel electrophoresis.

4) VNTRO are multiplied through PCR technique.

5) The separated fragments of DNA in the gel are copied on to a nylon

paper by southern blotting technique.

6) Special DNA probes are made in the laboratory. DNA-probes contain

repeated sequenced of bases complementary to those on VNTRs.

These probes are made radioactive by labeling with radioactive

isotopes. The radioactive DNA-probes bind to the repeat sequences on

the nylon membrane.

7) The dark bands on X-ray film represent the DNA fingerprints (=DNA

profiles).
 APPLICATIONS OF DNA FINGERPRINTING:

1. Paternity and Maternity

Because a person inherits his or her VNTRs from his or her parents,

VNTR patterns can be used to establish paternity and maternity. The

patterns are so specific that a parental VNTR pattern can be reconstructed

even if only the children’s VNTR patterns are known (the more children

produced, the more reliable the reconstruction). 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.

2. 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.

3. 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. Social security numbers, picture ID, and other

more mundane methods are much more likely to remain the prevalent ways

to establish personal Identification.

 PROBLEMS WITH DNA FINGER PRINTING

Like nearly everything else in the scientific world, nothing about

DNA fingerprinting is 100% assured. The term DNA fingerprint is, in one

sense, a misnomer: it implies that, like a fingerprint, the VNTR pattern for a

given person is utterly and completely unique to that person. Actually, all

that a VNTR pattern can do is present a probability that the person in

question is indeed the person to whom the VNTR pattern (of the child, the

criminal evidence, or whatever else) belongs. Given, that probability might

be 1 in 20 billion, which would indicate that the person can be reasonably

matched with the DNA fingerprint then again, that probability might only be

1 in 20, leaving a large amount of doubt regarding the specific identity of the

VNTR pattern’s owner.

1. Generating a High Probability

The probability of a DNA fingerprint belonging to a specific person

needs to be reasonably high-especially in criminal cases, where the

association helps establish a suspect’s guilt or innocence. Using certain rare

VNTRs or combinations of VNTRs to create the VNTR pattern increases the

probability that the two DNA samples do indeed match (as apposed to look
alike, but not actually come from the same person) or correlate (in the case

of parents and children).

2. Problems with Determining Probability

a. Population Genetics

VNTRs, because they are results of genetic inheritance, are not

distributed evenly across all of human population. A given VNTR cannot,

therefore, have a stable probability of occurrence; it will vary depending on

an individual’s genetic background. The difference in probabilities is

particularly visible across racial lines. Some VNTRs that occur very

frequently among Hispanics will occur very rarely among Caucasians or

African-Americans. Currently, not enough is known about the VNTR

frequency distributions among ethnic groups to determine accurate

probabilities for individuals within those groups; the heterogeneous genetic

composition of interracial individuals, who are growing in camber, presents

an entirely new set of questions. Further experimentation in this area, known

as population genetics, as been surrounded with and hindered by

controversy, because the idea of identifying people through genetic

anomalies along racial lines comes alarmingly close to the eugenics and

ethnic purification movements of the recent past, and, some argue, could

provide a scientific basis for racial discrimination.

B. Technical Difficulties

Errors in the hybridization and probing process must also be figured

into the probability, and often the idea of error is simply not acceptable.

Most people will agree that an innocent person should not be sent to jail, a

guilty person allowed to walk free, or a biological mother denied her legal

right to custody of her children, simply because a lab technician did not

conduct an experiment accurately. When the DNA sample available is

minuscule, this is an important consideration, because there is not much

room for error, especially if the analysis of the DNA sample involves

amplification of the sample (creating a much larger sample of genetically

identical DNA from what little material is available), because if the wrong

DNA is amplified (i.e. a skin cell from the lab technician) the consequences

can be profoundly detrimental. Until recently, the standards for determining

DNA fingerprinting matches, and for laboratory security and accuracy which

would minimize error, were neither stringent nor universally codified,

causing a great deal public outcry.