The techniques of medical science applied to assist in the resolution of crimes, legal disputes etc. constitutes forensic medicine. Establishing the identity of victim (e.g of murder, accidents etc) is critical to solving the problems of crimes cases. DNA fingerprinting or DNA profiling, is a highly sensitive and extremely versatile approach to this problem.
Preparation of DNA sample: Common sources of DNA include blood, sermen solid tissures blood stains, semen stains, hair roots etc. The specimens should be stored under clean, dry, cool condition to minimize contamination by microorganisms and DNA degradation due to DNase .
The DNA is prepared as follows.
Cells lysed with the enzyme proteinase K which also digests protein residual protein may be separated using the detergent sodium dodecyl sulphate RNA can be eliminated by phenol pure high molecules weight DNA may be prepared using alternative protocol.
In case of mixed specimens, suitable separation procedures should be used in separate the constituent cells. For example viginal swabs from rape types can be extracted separately following specific procedures.
The degree of DNA degradation is assessed as follows soutnern blottin is combined with species specific probes to establish DNA identity of human origin or otherwise DNA degradation is estimated by electrophoresis in agarose gel staining with ethidium bromide and observation under UV light high molecular weight DNA appears as single large band whereas degraded DNA forms a large smear.
DNA quantity in the sample is determined by absorbance 260 nm agarose gel electrophoresis with DNA preparations of known quantity and staining with ethidium bromide to assess the intensity of stain taken and fixing the DNA on membrane filter and hybridizing with a labelled highly repetitive primate specific locus.
Approach of DNA analysis: Many polymorphic loci composed of tandem repeats of 2-80 base pair the number of repeats varying from 1-100 VNTR and polymorphic for the number of tandem repeats while minivariant repeats show polymorphism for base pair composition at specific sites within each repeat. This variation at one or more loci may be analyzed by RFLP or PCR can also be used to analyze polymorphic site that may or may not be located in tandem repeat units. The DNA sample or the PCR products may be analyzed by the following approaches.
VNTR analysis: The sample DNA is used for RFLP analysis this requires high molecular weight DNA PCR products are used to develop RAPD pattern this can use much smaller even somewhat degraded DNA preparations.
Dot blot Analysis: The DNA samples is subjected to PCR to amplify the loci polymorphism at which is be determined. The PCR products are fixed to membrane and hybridized with allele specific probes.
Base sequence Analysis: The base sequence of specific loci in the region of DNA can be determined variable region by DNA sequencing and used for preparing DNA profiles.