o a FAM, 6-carboxyfluorescein; NED, 2'-chloro-5'-fluoro-7',8'-fused phenyl-1,4-dichloro-6-carboxyfluorescein; VIC, 2'-chloro-7'-phenyl-1,4-dichloro-6-carboxyfluorescein.
is relatively conserved among different species and perhaps it is not sufficiently primate-specific to be of general forensic utility (Timken et al., 2005).
A TaqMan-MGB (minus groove binder) real-time PCR design has been developed to target a small region of 62 bp located 31 bp downstream from the polymorphic repeat region of the HumTH01 locus. The assay has been applied to the quantification of human nuDNA from a variety of body fluid stains (Richard et al., 2003).
A human DNA quantification kit was developed for the quantification of human nuDNA by targeting a 62 bp portion of the human telomerase reverse transcriptase (hTERT) locus using a TaqMan-MGB assay that includes an internal PCR control (IPC) for the assessment of PCR efficiency against Taq inhibitors (Green et al., 2005). The kit has been validated for use in forensic casework according to the Scientific Working Group on DNA Analysis Methods (SWGDAM) guidelines and now is the most used assay in forensic casework for nuDNA quantitation.
More recently, a multiplex quantitative PCR assay has been described to amplify simultaneously two target sequences of different length, the TH01 STR locus (170-190 bp) and the upstream flanking region of the CSF1PO STR locus (67 bp), which allows for the assessment of DNA degradation in samples of forensic interest (Swango et al., 2006). The assay also includes an internal PCR control target sequence to allow for an assessment of PCR inhibition.
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