FORENSIC DNA ANALYSIS
DNA can be collected from: Anything that contains cells.
Body Fluids: blood; saliva; semen; others.
Body Tissues: skin; hair; bones; organs.
DNA Polymorphisms: A polymorphism is a gene or loci with two or more alleles of appreciable frequency. For the purposes of forensics, DNA polymorphisms are specific sites in the genome where the precise sequence of DNA tends to differ in unrelated individuals. These polymorphisms are sometimes found in genes, accounting for the differences in phenotype that we observe between individuals. Within genes, polymorphisms are usually referred to as "variants". There is evolutionary pressure exerted on polymorphisms that occur within genes. In most cases, alteration of DNA sequence will result in the alteration of the function of a particular protein (deleterious mutation). This alteration may make the individual either more or less fit to survive. Evolution thus limits the number of polymorphisms that accumulate within functional genes (exons).
Natural selection does not operate in this way on inter-gene ("between the genes") regions (introns) of the genome. Introns have been referred to as "junk DNA" because their code is not read and translated into functional proteins. These regions are large in most eucaryotic organisms: in the human genome more than 50% of the DNA seems not to be associated with any gene. Because there is no selective pressure against introns, large numbers of polymorphisms have accumulated in these non-coding regions of DNA. These polymorphisms have turned out to be valuable for genetic mapping and forensic identification.
Polymorphism Types: There are several types of DNA polymorphisms that tend to be useful for genetic mapping and forensic analysis:
RFLP: RFLP stands for restriction fragment length polymorphism. Simple RFLP's are single base pair alterations in the DNA sequence at a particular locus on the genome. The alteration in sequence is such that the recognition site for a particular restriction enzyme is either added or eliminated. Thus the pattern of restriction fragments or cuts generated by digestion of DNA with the enzyme (DNA polymerase) in question will vary.
VNTR: VNTR is the abbreviation for Variable Number of Tandem Repeats. VNTRs are polymorphisms where a particular sequence is repeated at that locus numerous times.
TATTTATTTATTTATTTATTTATTTATTTATTTATTTATTTATTTATT
Short Tandem Repeats (STR) are repetitive sequences that are usually small (less than 20 base pairs). The number of times a sequence is repeated can vary from just a few to up to 60 or more. VNTRs are detected as variations in fragment length between two fixed points outside of the VNTR region..
DNA Profiles from 12 Unrelated Individuals at a Polymorphic Locus
VNTRs are highly variable when compared with simple RFLPs-- many different patterns are possible with VNTRs, whereas with simple RFLPs there are only two possibilities, either a restriction site is there, or its not. VNTRs are thus more useful for forensic applications.
Interspersed Repetitive DNA. Interspersed repetitive DNA is a type of polymorphism where a particular block is repeated, not sequentially at a single locus (as was the case with VNTRs), but rather throughout the genome. Only one copy of the sequence is observed at any given locus, but that same block of sequence is observed at thousands of different loci.
Interspersed repetitive DNA is thought to make up between 5% - 20% of the human genome. The percentages, specific distribution patterns, sizes, and sequences of the repetitive blocks are generally consistent within a species, but vary between species. For this reason, interspersed repetitive DNA sequences have been useful in tracing evolutionary relationships between species. Interspersed repetitive DNA elements are also referred to as "satellite DNA" because they are usually located adjacent to coding regions of the DNA (genes).
Polymerase Chain Reaction (PCR)
PCR involves the repeated copying - known as PCR amplification - of specified areas of DNA molecules. These areas are known as alleles and are identified by specific sequences of bases at each of their ends known as flankers. The flankers occur at the same locations on the chromosomes of all people. With each copying cycle, the quantity of the target allele is doubled - after 30 copying cycles there will be one billion times more copies of the target alleles than at the start of the amplification process.
Depending on the loci examined, analysis of variation is carried out using one of two approaches: sequence variation or length variation. Sequence variation looks at variation in the sequence of bases. If the target sequence is thought of as the spelling of a word, then sequence variation identifies the particular spelling of that word for an individual.
METHODS: Restriction Fragment Length Polymorphism used to detect VNTR sequences. This method is being replaced by PCR of STRs.