The technique of DNA fingerprinting and its applications


Analysis of diverse and polymorphic DNA sequences is called DNA fingerprinting. DNA fingerprinting is not only useful in solving parental disputes but can be used in the identification of victims of accidents and criminals. Go through this article and get to know about the technique and applications of DNA fingerprinting.

Introduction


The analysis of diverse polymorphic DNA sequences of DNA sample is called DNA fingerprinting. DNA fingerprinting is based on restriction fragment length polymorphism RFLP, polymerase chain reaction PCR, dot blot assay and determination of base sequence. Alec Jeffrey 1985 first of all developed this technique. DNA fingerprinting is not only useful in solving parental disputes but it is also an important tool in criminal investigations.

In human DNA, few sequences of nucleotides repeatedly occur. These sequences may be many bases long. The frequency of these sequences is different in different individuals. The hypervariability of DNA depends on the number of nucleotides present in these areas and their repetition. These areas are called variable number tandem repeat VNTR loci. Microsatellites and minisatellites are included in VNTR. Microsatellite repeats are 1-6 bp long while minisatellites are up to 1 kbps long. VNTRs are used in DNA fingerprinting.

Technique of DNA Fingerprinting


For DNA fingerprinting about 1 microgram DNA is required which can be obtained from blood, skin cells, semen or hair follicles. The process of DNA fingerprinting can be divided into the following steps:

  1. Isolation of DNA: IN the first step, to isolate DNA from the tissue, the cells are lysed with an appropriate detergent chemical like SDS. The proteins obtained from lysis are digested with proteinase K enzyme in the presence of EDTA.
    Now DNA is extracted with the help of phenol. Nowadays DNA is extracted from the cells by high-speed refrigerated centrifuge. If the DNA is present in limited amount then it is amplified by making many copies of it by a polymerase chain reaction.

  2. Digestion of DNA with restriction enzyme : After the extraction of DNA it is broken into pieces with the help of some restriction enzymes . The fragments thus formed have the same type of terminal nucleotide sequence.

  3. Gel electrophoresis of DNA: In gel electrophoresis DNA fragments are separated on the basis of their size and weight. For this high voltage electric current is used to separate DNA fragments. The mixture of DNA fragments is filled in the wells made in agarose gel. Is negatively charged molecules, on passing electric current its fragments disperse with different speed on the basis of their size and weight. The dispersed fragments of DNA from different bands can be seen in ultraviolet light.

  4. Transfer of DNA on Nylon membrane : DNA is a double-stranded molecule hence alkaline chemicals are used to separate its strands. After its reaction with alkaline chemicals, the gel is placed on filter paper dipped in an appropriate buffer. Now nitrocellulose filter is placed on the gel and covered with a stack of filter papers. Due to capillary action buffer moves from the dipped filter paper to the dry stack of filter papers through the gel and nitrocellulose membrane. DNA fragments also move upwards along with buffer and get entangled in nitrocellulose filter. This process is known as Southern blotting technique after the name of its inventor E.M. Southern. This nitrocellulose filter is baked at 80 degree Celsius, therefore, the DNA molecules get stuck to it permanently.

  5. Radiolabelling of DNA : To show the DNA fragments present on nitrocellulose membrane this membrane is dipped in a solution which already has labeled radioactive DNA fragments as probes. These probes identify a particular nucleotide segment which is complementary to the VNTRs. This process is known as hybridization. Now the gel is washed repeatedly to remove the hybridized probes. This membrane is exposed to an X-ray film and the film is developed which shows bands at such places where the complementary sequence of the probe might have been present on the DNA segment. The banding pattern of DNA is called fingerprint.


Applications of DNA fingerprinting


The applications of DNA fingerprinting are as follows:

  • DNA fingerprinting helps in the study and understanding of the evolutionary distance between two adjacent groups in the evolutionary tree.

  • In Forensic science, DNA fingerprinting is used to solve problems of rape and murder cases, paternity, etc.

  • DNA fingerprinting is used in the identification of recombinant DNA.

  • In the chemical field, DNA fingerprinting is used in the diagnosis of pathogens.


Comments

Author: Umesh23 Apr 2019 Member Level: Diamond   Points : 2

It is a highly technical writeup presented by the author in a simple language. Overall an informative article about DNA fingerprinting. Some of the abbreviations and units are not very clear and it would have been better if the author had given their full forms.

DNA fingerprinting has evolved as a modern method in various areas including the forensic investigations which are crucial in unearthing the truth in complicated cases. Just as two fingerprints are not similar in the same way, the two DNA patterns are also not similar. So this has been a great scientific method for identification of the person whose blood samples are got for analysis.



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