Mitochondrial DNA, Y-STRs and emerging forensic DNA tools (Rebecca Just, Armed Forces DNA Identification Laboratory)

The mitochondrial genome (mtGenome), a circular, ~16,000bp genome located in the mitochondria of cells, and the Y-chromosome are two markers that are inherited uniparentally; the mtGenome is inherited strictly from the mother, and the Y-chromosome only from the father. As the mtGenome and a large portion of the Y-chromosome are inherited without recombination, these markers are often used to reconstruct human migration, to examine population admixture, and to make determinations about biogeographic ancestry. These markers can also have substantial utility in forensics depending on the type of biological evidence and genetic references available.

Due to its high copy number (~1000 copies per cell), mtDNA is often examined in situations where the DNA is highly degraded or the sample is unlikely to have much nuclear DNA (ie. hair shafts or fingernails). mtDNA is most typically examined by Sanger sequencing of the two hypervariable regions or the entire control (non-coding) region.

As the sex-determining Y-chromosome is generally present only in males, Y-STRs are often examined in cases where it is necessary to examine a male genetic profile separate from the female component of a mixed sample (ie. a rape case). Y-STRs are typed in the same manner as autosomal STRs, and several commercial kits are available.

While a strength in some respects, the uniparental inheritance pattern and lack of recombination of mtDNA and the Y-chromosome necessitates that a “counting method”, rather than the product rule, be used to evaluate the statistical strength of such genetic evidence in a case. Additionally, the high mutation rate of both markers systems requires more conservative “match” interpretation.

In addition to mtDNA sequencing, other tools now exist for typing degraded human samples. One recent development is “miniSTRs”: standard STR markers with the primer binding sites designed closer to the repeat regions. This design strategy results in shorter amplicons, and thus often more successful typing on highly fragmented DNA. The small size of the amplicons, however, limits the available real estate within a multiplex, which results in fewer markers per multiplex and a lower power of discrimination overall.

Though less typically applied, single nucleotide polymorphisms (SNPs) can also be used to type degraded samples. Because each SNP involves only a single base position, small amplicons are both practical and typical. Nuclear SNPs have a much lower power of discrimination than STRs, however many SNP typing and detection methods allow for the construction of large multiplex panels, enabling the typing of many SNPs at once. mtDNA SNPs can also be used to improve upon the discrimination obtained from control region sequencing alone.

Materials

AFDIL policy prohibits the publishing of presentations

mtDNA

Holland and Parsons. Mitochondrial DNA sequence analysis – validation and use for forensic casework. Forensic Sci. Rev. 1999;11:22-50.

Carracedo et al. DNA Commission of the International Society for Forensic Genetics: guidelines for mitochondrial DNA typing. Forensic Sci. Intl. 2000;110:79-85.

Gabriel et al. Improved mtDNA sequence analysis of forensic remains using a “mini primer set” amplification strategy. J. Forensic Sci. 2001;46:247-253.

Melton and Nelson. Forensic mitochondrial DNA analysis: two years of commercial casework experience in the United States. Croatian Med. J. 2001;42(3):298-303.

Y-STRs

Kayser and Sajantila. Mutations at Y-STR loci: implications for paternity and forensic analysis. Forensic Sci. Intl. 2001;118:116-121.

Mulero et al. Development and validation of the AmpFlSTR® Y-Filer™ PCR amplification kit: A male-specific, single amplification 17 Y-STR multiplex system. J. Forensic Sci. 2006;51(1):64-75.

Gill, et al. DNA Commission of the International Society of Forensic Genetics: recommendations on forensic analysis using Y-chromosome STRs. Int. J. Legal Med. 2001;114(6):305-309.

Mini-STRs

Wiegand and Kleiber. Less is more - Length reduction of STR amplicons using redesigned primers. Int. J. Leg. Med. 2001;114(4-5):285-287.

Butler, Shen and McCord. The development of reduced size STR amplicons as tools for analysis of degraded DNA. J. Forensic Sci. 2003;485(5):1054-1064.

SNPs

Coble et al. Single nucleotide polymorphisms over the mtDNA genome that increase the power of forensic testing in Caucasians. Int. J. Leg. Med. 2004;118:137-146.

Vallone et al. A multiplex allele-specific primer extension assay for forensically informative SNPs distributed throughout the mitochondrial genome. Int. J. Leg. Med. 2004;118:147-57.

Gill et al. An assessment of whether SNPs will replace STRs in national DNA databases – joint considerations of the DNA working group of the European Network of Forensic Science Institutes (ENFSI) and the Scientific Working Group on DNA Analysis Methods (SWGDAM). Sci. Justice. 2004;44:51–53.