DNA & genetics
When nuclear DNA fails: mitochondrial DNA analysis for degraded and burned remains
Why mitochondrial DNA survives in bone, teeth and hair when STR profiling fails, how laboratories sequence it, and what a mitochondrial match can and cannot prove.
Forensic Science Pakistan3 min read
Standard STR profiling reads nuclear DNA, which exists as just two copies in each cell. Fire, decades in the ground or a damp grave break those molecules into fragments too short to amplify, and the profile drops out locus by locus. When that happens, laboratories turn to mitochondrial DNA (mtDNA).
Why mitochondrial DNA survives
- Copy number: a cell holds hundreds to thousands of copies of the mitochondrial genome against two of the nuclear genome, so some intact copies usually remain even after heavy damage.
- Size: the genome is a small circle of 16,569 base pairs.
- Where it is found: dense bone, tooth dentine and hair shafts without roots, samples that often yield no nuclear profile at all.
The control region and its hypervariable segments
Forensic sequencing concentrates on the non-coding control region, about 1,122 base pairs from position 16024 to 576, which crosses position 0 of the circular genome. Most of the variation between people sits in three hypervariable segments: HV1 (16024–16365), HV2 (73–340) and HV3 (438–574).
Results are written as differences from the revised Cambridge Reference Sequence (rCRS). The reference happens to carry a rare adenine at position 263, so most people are reported as 263G. An extra cytosine inserted after position 315 is written 315.1C.
From bone to sequence
- Clean the surface: abrade the outer bone and treat it with bleach and UV light to remove modern DNA.
- Powder: drill or mill a small sample of dense bone or tooth into fine powder.
- Demineralise and digest: dissolve the mineral with EDTA and digest proteins with proteinase K to release DNA.
- Purify: bind DNA to silica to separate it from calcium, soil humic acids and other PCR inhibitors.
- Amplify short targets: when DNA is badly fragmented, overlapping mini-amplicons of around 100 base pairs replace the usual longer ones.
- Sequence: Sanger sequencing reads both strands; massively parallel sequencing can read the whole mitochondrial genome.
For the oldest and most damaged remains, the best sample is often the inner ear part of the skull; see why the petrous bone is the gold standard.
Heteroplasmy
Some people carry more than one mtDNA sequence. Point heteroplasmy shows as two overlapping bases at one position. Length heteroplasmy appears in runs of cytosines, such as the C-stretch around 16184–16193 in HV1, where the number of Cs varies between molecules. Because of this, interpretation guidelines generally treat a single difference between an evidence sample and a reference as inconclusive rather than an exclusion; two or more clear differences exclude.
Contamination
The same sensitivity that makes mtDNA useful makes it easy to contaminate. Laboratories work in separate pre- and post-PCR areas, run blank extractions and negative controls with every batch, and keep staff profiles to eliminate their own DNA, the same precautions used in ancient DNA clean rooms. An unbroken chain of custody matters just as much.
What a mitochondrial match means
Mitochondrial DNA passes from a mother to all her children without recombining, so everyone in a maternal line shares the same sequence. That has two consequences:
- Any maternal relative can be the reference: a missing person’s mother, siblings, maternal aunts and uncles, or the children of maternal aunts. That makes it valuable for identifying the missing and disaster victims.
- It cannot single out one person: a match means the sample came from that person or someone in their maternal line.
For the same reason, a mitochondrial match is not reported as a random match probability. Laboratories search a population database such as EMPOP and report how often the sequence has been seen, which depends on having good reference data for the population concerned. For relationship cases built on nuclear DNA, see our guide to the DNA test in Pakistan.
