99.9 per cent of human base sequences are same. The difference of 0.01 percent in the DNA sequence makes different individual unique in their phenotypic appearances. To find out the genetic differences between two individuals or among individuals of a population, sequencing the DNA every time would be a tedious and expensive task. DNA fingerprinting is a very quick way to compare the DNA sequences of any two individuals. It is a method of isolation and identification of variable elements within the base-pair sequence of DNA. It is also known as DNA profiling, DNA typing, genotyping or genetic fingerprinting.
DNA fingerprinting involves the identification of differences in some specific regions in DNA sequences which are known as repetitive DNA. In these DNA sequences, a small stretch of DNA is repeated many times.
During density gradient centrifugation, these repetitive DNA are separated from bulk genomic DNA as they form different peaks. Major peak is formed by bulk DNA and the other small peaks are referred as satellite DNA. Depending on number of repetitive units, base composition (A:T rich or G:C rich) and length of segment; the satellite DNA is classified into many categories such as mini-satellite, micro-satellites, etc. Generally, these sequences do not code for any protein, but they form a large portion of human genome. These satellite DNA sequences exhibit a high degree of polymorphism and thus form the basis of DNA fingerprinting. DNA from all tissues (like hair-follicle, sperm, skin, saliva, bone, blood, etc.) from an individual shows the same degree of polymorphism, thus it becomes an important tool in forensic application. Polymorphism is inheritable from parents to children, thus DNA fingerprinting is used in paternity testing.
Polymorphism is the variation at genetic level which arises due to mutations. Mutations can arise in an individual either in somatic cells or in the germ cells. If an inheritable mutation occurs in human population at high frequency, it is termed as DNA polymorphism. These mutations keep on accumulating generation after generation and form one of the basis of polymorphism. There are different types of polymorphism ranging from one single nucleotide to very large scale. Such polymorphisms play an important role in speciation and evolution.
DEVELOPMENT OF DNA FINGERPRINTING
DNA Fingerprinting was initially developed by Alec Jeffreys. A satellite DNA which shows a very high degree of polymorphism was used as probe by him. It is known as Variable Number of Tandem Repeats. DNA fingerprinting technique involves the following steps:
- Isolation of DNA
- Digestion of DNA by restriction endonucleases
- Separation of DNA fragments by electrophoresis
- Transfer of separated DNA fragments to synthetic membranes, such as nylon or nitrocellulose. This is known as blotting.
- Hybridisation using radio labelled Variable Number of Tandem Repeat (VNTR) probe
- Detection of hybridised DNA fragments by autoradiography
The VNTR belongs to a mini-satellite category of satellite DNA. A small DNA sequence is tandemly arranged in many copy numbers. In an individual, the copy number varies from chromosome to chromosome. Degree of polymorphism depends upon the number of repeatations. As a result, the size of VNTR varies from 0.1 to 20 kb. After the hybridisation with VNTR probe, autoradiogram shows many bands of different sizes. These bands show a characteristic pattern for an individual DNA. This pattern is different for every 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. Thus, DNA from a single cell is enough to perform DNA fingerprinting analysis.
APPLICATIONS OF DNA FINGERPRINTING
- It is used to determine the father or a mother of a child. VNTR is inheritable. Thus, if child's VNTR is known, the parental VNTR pattern can be recreated. It can be used to determine the biological father in identification, adoption, confirming legal nationality and biological parenthood cases.
- DNA taken from skin, hair cells, blood or any other DNA evidence left at the scene of crime can be compared through the banding patterns of DNA fingerprinting. By comparing it with the DNA of the suspect, the authorities can determine that whether the suspect is guilty or innocent. It can also be used to determine the identity of a homicide victim by the similar manner.
- It helps the doctors and scientists to find out the best possible match in case of an organ transplant operation.
- It helps to identify victims in case of disaster.
- It helps in studying the variation within a population or ethnic group.
- It is used to identify the inherited disorders (like Huntington’s disease, sickle cell anemia, cystic fibrosis, etc.) in unborn babies, children and adults.
- It is used to trace ancestors and create a family tree.
- It is used in studying the genetic variability of endangered species in conservation biology.
- It is used to test the pathogens in food.