11-10-2012, 03:04 AM
(This post was last modified: 11-10-2012, 03:51 AM by Administrator.)
Application of knowledge of DNA based information for resolving crimes and knowing the identity of criminals is known as forensic science. DNA fingerprinting is a highly accurate and sensitive mode of approach of resolution of crimes.
The technique of DNA fingerprinting was first demonstrated by Alec Jeffreys in 1986.
Principle behind the DNA fingerprinting or profiling process is to determine and indicate the presence of specific allele in a series of polymorphic loci or locus in his/her genome. A polymorphic locus refers to a region in the genome of an individual different from others. The presence of different alleles in such a locus is studied by using restricted fragment length polymorphism (RFLP) or by PCR amplification. Thus, the whole number of polymorphism existing in the mini, micro satellite and mini variant repeats constitute the variable number of tandem repeats(VNTR) and are analysed for the DNA fingerprinting process. The site of presence of VNTR is known as VNTR locus. The different allele consists of different number of repeating units of VNTR sequence. The sequence of unique DNA elements flanking the repeats are known and primers corresponding to the same is produced and used for amplification in PCR of mini and micro satellite DNA. The final product of such amplification tends to be different in different individual and can be separated by gel electrophoresis technique.
The DNA is extracted from several sources like hair, semen, solid tissues, blood tissues, blood stains, buccal cells of saliva etc. The extracted DNA is stored in a clean, cool, dry place. In the cells extracted, it may be digested by sodium do decyl sulphate, and the proteins digested by some proteinase enzyme. This is followed by repeated extraction of these with phenol and chloroform to remove cell debris. The obtained DNA is dissolved in a solution. This can be precipitated by addition of alcohol. DNA degradation is specified by staining DNA with ethidium bromide and undergoing agarose gel electrophoresis. The denatured DNA appears as smear whereas those not digested appears as a single band.
DNA profiling
The DNA obtained is studied by mainly two mechanisms –by RFLP and by PCR analysis.
RFLP: - These analyses are of two types: multi locus polymorphism (MLP) and single locus polymorphism (SLP) .The DNA digested and run on agarose is transferred to a membrane and probed with highly specified probe. Thus probe also hybridize with fragments of similar sequences. The hybridized probe is detected by autoradiography or in the case of alkaline phosphatase present in the probe by chemiluminescence. It is not possible to differentiate fragments differing by few repeats by this process.
PCR: This analysis are of two types STR (short tandem repeats) and MVR (minisatellite variant repeat) analyses. The most common type is to analyse STR. In this methods two probes complementary to the sequences flanking an STR is used to hybridize and each STR obtained is amplified by PCR. The resulting amplified product appears as distinct bands in agarose gel. Each STR has as many as 9 to 10 different alleles in human. Each individual has two loci for STR. Therefore it is necessary to analyse 9 in 10 repeats to attain a definite result. This process has several advantages like the procedure is very sensitive and DNA can be retrieved from very minute specimens available, the result is obtained very quickly and even the fragmented DNA can be used as primary sample for this purpose. Thus a single PCR cycle provides us with multiple copies to work on.
The information of the DNA obtained by such methods is used to interpret the results of the crime. For eg: the band of DNA is compared with those of the DNA obtained at the crime spot. If the bands are similar then the accused can be confirmed as the culprit. Thus it enables a high sensitive technique for identification of the criminals.
The advantages of DNA profiling include
Identification of criminals: The main and most widely used application of DNA profiling is use of the technology to identify criminal. This helps in determining whether the accused is the real culprit or not.
Kinship analysis: This method can be use to determine where two or more individual are members of the same family. It is also an important evidence for paternity testing in confirming the parents of a child.
Sex identification: This can also be used for identification of sex by amplifying the sequence for Y chromosome.
The technique of DNA fingerprinting was first demonstrated by Alec Jeffreys in 1986.
Principle behind the DNA fingerprinting or profiling process is to determine and indicate the presence of specific allele in a series of polymorphic loci or locus in his/her genome. A polymorphic locus refers to a region in the genome of an individual different from others. The presence of different alleles in such a locus is studied by using restricted fragment length polymorphism (RFLP) or by PCR amplification. Thus, the whole number of polymorphism existing in the mini, micro satellite and mini variant repeats constitute the variable number of tandem repeats(VNTR) and are analysed for the DNA fingerprinting process. The site of presence of VNTR is known as VNTR locus. The different allele consists of different number of repeating units of VNTR sequence. The sequence of unique DNA elements flanking the repeats are known and primers corresponding to the same is produced and used for amplification in PCR of mini and micro satellite DNA. The final product of such amplification tends to be different in different individual and can be separated by gel electrophoresis technique.
The DNA is extracted from several sources like hair, semen, solid tissues, blood tissues, blood stains, buccal cells of saliva etc. The extracted DNA is stored in a clean, cool, dry place. In the cells extracted, it may be digested by sodium do decyl sulphate, and the proteins digested by some proteinase enzyme. This is followed by repeated extraction of these with phenol and chloroform to remove cell debris. The obtained DNA is dissolved in a solution. This can be precipitated by addition of alcohol. DNA degradation is specified by staining DNA with ethidium bromide and undergoing agarose gel electrophoresis. The denatured DNA appears as smear whereas those not digested appears as a single band.
DNA profiling
The DNA obtained is studied by mainly two mechanisms –by RFLP and by PCR analysis.
RFLP: - These analyses are of two types: multi locus polymorphism (MLP) and single locus polymorphism (SLP) .The DNA digested and run on agarose is transferred to a membrane and probed with highly specified probe. Thus probe also hybridize with fragments of similar sequences. The hybridized probe is detected by autoradiography or in the case of alkaline phosphatase present in the probe by chemiluminescence. It is not possible to differentiate fragments differing by few repeats by this process.
PCR: This analysis are of two types STR (short tandem repeats) and MVR (minisatellite variant repeat) analyses. The most common type is to analyse STR. In this methods two probes complementary to the sequences flanking an STR is used to hybridize and each STR obtained is amplified by PCR. The resulting amplified product appears as distinct bands in agarose gel. Each STR has as many as 9 to 10 different alleles in human. Each individual has two loci for STR. Therefore it is necessary to analyse 9 in 10 repeats to attain a definite result. This process has several advantages like the procedure is very sensitive and DNA can be retrieved from very minute specimens available, the result is obtained very quickly and even the fragmented DNA can be used as primary sample for this purpose. Thus a single PCR cycle provides us with multiple copies to work on.
The information of the DNA obtained by such methods is used to interpret the results of the crime. For eg: the band of DNA is compared with those of the DNA obtained at the crime spot. If the bands are similar then the accused can be confirmed as the culprit. Thus it enables a high sensitive technique for identification of the criminals.
The advantages of DNA profiling include
Identification of criminals: The main and most widely used application of DNA profiling is use of the technology to identify criminal. This helps in determining whether the accused is the real culprit or not.
Kinship analysis: This method can be use to determine where two or more individual are members of the same family. It is also an important evidence for paternity testing in confirming the parents of a child.
Sex identification: This can also be used for identification of sex by amplifying the sequence for Y chromosome.
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