One of the major challenges of mtDNA typing lies in the fact that many sequences fall into common groupings termed 'most common types'. For example, a review of the HV1/HV2 type distribution in 1655 Caucasians of U.S. and European descent (Monson et al. 2002) found that the most common mtDNA
Samples of diverse worldwide origin
Samples of diverse worldwide origin
African, Asian, European origin
560 (coding region only)
Ingman et al. (2000) AF346963-AF347015
Maca-Meyer et al. (2001) AF381981-AF382013
Herrnstadt et al. (2002) Sequences available at www.mitokor.com
24 PCR reactions, 48 sequencing reactions
32 PCR reactions, 64 sequencing reactions
68 PCR reactions, 136 sequencing reactions
Summary of published mtGenome DNA sequencing efforts from December 2000 to February 2004 representing almost 1000 complete mtGenomes.
East Asian lineages 48
Australian and 52
New Guinean Aborigines and Polynesians
Most common 241
Kong et al. (2003) AY255133-AY255180
Ingman and Gyllensten (2003) AY289051-AY289102
Coble et al. (2004) AY495090-AY495330
15 PCR reactions,
47 sequencing reactions
24 PCR reactions,
12 PCR reactions, 95 sequencing reactions type, which matches the rCRS, occurred 7.1% of the time (Coble et al. 2004). Furthermore, it was observed that only 18 mtDNA types account for 20.8% of the total Caucasian data set (Coble et al. 2004). The presence of these most common types suggests that one of out every five times a mtDNA sequence analysis is performed on a Caucasian individual, the result would be expected to match numerous other individuals in a population database. While the same analysis revealed that approximately 50% of the 1655 individuals present in the European Caucasian population are 'unique in the database', having a sample that falls into one of these most common types can be present a disappointing statistic after all of the hard work taken to generate the full mtDNA HV1/HV2 sequence.
An extensive search for distinguishing single nucleotide polymorphisms in samples possessing the most common Caucasian types was recently undertaken (Parsons and Coble 2001). A total of 241 complete mtGenomes were sequenced from the 18 common European Caucasian HV1/HV2 types mentioned above (Coble 2004, Coble et al. 2004). The samples typed come from mtDNA haplogroups H, J, T, V, and K (see the next section for more discussion on haplogroups).
Examination of whole mtGenome sequence information expanded the 18 most common Caucasian HV1/HV2 types to 209 resolvable haplotypes (Coble et al. 2004). This almost 12-fold improvement in resolving power for these common HV1 /HV2 types required about 27 times the amount of DNA sequencing - from 610 bases for just HV1/HV2 alone to ~16 569 for the entire mtGenome. Obviously, this approach is not a cost effective one. Furthermore, even with the expansion in sequence information, 32 of the 241 individuals matched one or more individuals across the entire mtGenome.
From their extensive sequencing information, Coble et al. (2004) selected a battery of SNP markers to aid in resolving the most common Caucasian mtDNA HV1/HV2 types without the costly and time-consuming venture of having to sequence the entire mtGenome. A total of 59 informative SNPs were placed into eight multiplex panels (Coble et al. 2004). The first panel provides maximum resolution of the most common Caucasian HV1/HV2 mtDNA type (i.e., that matching CRS) and examines the following nucleotides spanning the mtGenome: 477, 3010, 4580, 4793, 5004, 7028, 7202, 10211, 12858, 14470, and 16519. Vallone et al. (2004) combined these 11 SNP sites into a multiplex allele-specific primer extension or 'SNaPshot' assay (see Chapter 8) that can reliably type a sample that contains only a few hundred copies of mtDNA.
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