Figure 7.9 A mixture of two individuals will lead to up to four peaks at each locus. The area between the dotted lines represents the zone where the minor component of the mixture can be interpreted. The lower dotted line represents 15% and 60% of the major peaks – below 15% is a zone where stutter peaks from the major alleles can occur and peaks, below 60% cannot be easily explained by peak imbalance. At this locus the major component can be interpreted as 13-15 while the minor component’s genotype is 16 – 20
Many samples that are collected from a crime scene may have been exposed to the environment for hours, days, or even longer if the crime scene has gone undetected. When DNA analysis is being used to identify human remains, the remains may be several years old before they are analysed or may have been exposed to severe envi- ronmental insult such as high temperatures. In all these circumstances the DNA in the
Figure 7.10 The profile was generated using the AmpFlSTR® Profiler Plus® kit from Applied Biosys- tems. The DNA was extracted from a bone recovered from a Scottish loch after approximately 30 years. The profile is typical of a degraded profile with a gradual reduction in the amount of product as the amplicons increase in size (see plate section for full-colour version of this figure) cellular material will not be in pristine condition and will have degraded. This leads to a characteristic DNA profile with over amplification of the smaller loci and the suc- cessful amplification declines with the size of the alleles. Figures 7.10 and 7.11 show two examples of degraded DNA sample; the first one is from a bone sample that had been in water for 30 years. The small loci have over amplified whereas the larger loci are barely detectable  – the decrease in amplification is gradual as the length of the alleles increases. In the second example an example of locus drop out can be seen, the first two blue loci, D3S1358 and vWA have amplified successfully but there is no FGA allele. This
Figure 7.11 The profile was generated using the AmpFlSTR BlueTM kit from Applied Biosystems. The DNA was extracted from muscle tissue recovered from a plane crash. The muscle had been subjected to high temperatures and the DNA was highly degraded – no amplification products were detected from the FGA locus. The size standard is also shown in by non-shaded peaks (see plate section for full-colour version of this figure)
profile is from human muscle tissue that had been exposed to high temperatures and has degraded to the extent that there is very little or no DNA that is 200 bp or longer . The interpretation of degraded profiles can be difficult and particular attention has to be taken when homozygous loci are detected – are they really homozygous and not heterozygous with one of the alleles having dropped out? When the levels are very low, if there is enough material the PCR is carried out in duplicate, as with LCN PCR, to minimize the possibility of generating an incorrect profile. To assist with the analysis of degraded DNA, a series of multiplexes have been developed with the primers positioned close to the core repeats of the STRs, thereby minimizing the lengths of the amplicons [24-28].
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