Heteroplasmy

Heteroplasmy is the presence of more than one mtDNA type in an individual (Melton 2004). Two or more mtDNA populations may occur between cells in an individual, within a single cell, or within a single mitochondrion. It is now thought that all individuals are heteroplasmic at some level - many below the limits of detection in DNA sequence analysis (Comas et al. 1995, Bendall et al. 1996, Steighner et al. 1999, Tully et al. 2000). It is highly unlikely that millions of mtDNA molecules scattered throughout an individual's cells are completely identical given that regions of the mtGenome have been reported to evolve at 6-17 times the rate of single copy nuclear genes (see Brown et al. 1979, Wallace et al. 1987, Tully 1999). Consider that whereas only a single copy of each nuclear chromosome is present in an egg there are approximately 100 000 copies of the mtDNA genome present (Chen et al. 1995). Thus, for the transmission of a mtDNA mutation to become detectable it must spread to an appreciable frequency among a cell's mtDNA molecules.

Heteroplasmy may be observed in several ways: (1) individuals may have more than one mtDNA type in a single tissue; (2) individuals may exhibit one mtDNA type in one tissue and a different type in another tissue; and/or (3) individuals may be heteroplasmic in one tissue sample and homoplasmic in another tissue sample (Carracedo et al. 2000). Given that heteroplasmy happens, interpretation guidelines must take into account how to handle differences between known and questioned samples.

Both sequence and length heteroplasmy have been reported in the literature (Bendall and Sykes 1995, Bendall et al. 1996, Melton 2004). Length hetero-plasmies often occur around the homopolymeric C-stretches in HV1 at positions 16184-16193 and HV2 at positions 303-310 (Stewart et al. 2001) (see Figure 10.6). Sequence heteroplasmy is typically detected by the presence of two nucleotides at a single site, which show up as overlapping peaks in a sequence electropherogram (Figure 10.9).

Heteroplasmy at two sites in the same individual, a condition known as 'triplasmy', has been reported (Tully et al. 2000), but occurs at lower frequencies than single site heteroplasmy. Since it is rare to find more than one heteroplasmic position in the 610 nucleotides sequenced for HV1 and HV2, a report of as many as six heteroplasmic sites in an individual mtDNA sequence (Grzybowski 2000) raised suspicions about the sequencing strategy used. The Grybowski study has been criticized as possibly containing contamination due to the excessive number of amplification cycles used (Budowle et al. 2002a, Brandstätter and Parson 2003). A re-analysis of the same samples used in the original Grybowski study with a direct rather than a nested PCR approach resulted in a reduction in the reported number of samples with heteroplasmic positions (Grzybowski et al. 2003).

One of the major challenges of heteroplasmic samples is that the ratio of bases may not stay the same across different tissues, such as blood and hair or

Figure 10.9

(a) Sequence heteroplasmy at position 16093 possessing both C and

T nucleotides compared to

(b) the same region (positions 16086-16101) on a different sample containing only a T at position 16093.

Figure 10.9

(a) Sequence heteroplasmy at position 16093 possessing both C and

T nucleotides compared to

(b) the same region (positions 16086-16101) on a different sample containing only a T at position 16093.

between multiple hairs (Sullivan et al. 1997, Wilson et al. 1997, Sekiguchi et al. 2003). Some mtDNA protocols now recommend sequencing multiple hairs from an individual in order to confirm heteroplasmy.

Hotspots for heteroplasmy include the following positions in HV1: 16093, 16129, 16153, 16189, 16192, 16293, 16309, and 16337 (Stoneking 2000, Tully et al. 2000, Brandstätter and Parson 2003) and 72, 152, 189, 207, and 279 in HV2 (Calloway et al. 2000, Melton and Nelson 2001). One study found that the frequency of heteroplasmy can differ across tissue types with muscle tissue being the highest and was statistically significant across different age groups suggesting that heteroplasmy increases with age (Calloway et al. 2000). Heteroplasmy has also been reported to remain stable over time in the same individuals and thus be inherited rather than age related (Lagerström-Fermer et al. 2001). While heteroplasmy can sometimes complicate the interpretation of mtDNA results, the presence of heteroplasmy at identical sites can improve the probability of a match, such as seen in the Romanov study (see D.N.A. Box 10.2).

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  • ferumbras
    What is length heteroplasmy in forensic mtdna?
    2 months ago

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