M5 Flashcards
Methods used to ID human remains
- Facial characteristics
- Scars, marks, & other special body features
- Dental matching
- Fingerprint comparison
DNA profiles from human remains can be essential in:
- Missing person investigations
- Unidentified Remains Identification
- Mass disaster identification
If a reference sample is available, follows same underlying workflow of regular DNA profile as follows:
- Generate DNA profile from unknown evidentiary sample & known reference sample, or enter into CODIS
- Followed by comparison of the 2
- Inclusion/Exclusion/Incomplete
Generating DNA profiles from skeletonized/decomposed human remains is
challenging
NamUS
National Missing and Unidentified Person System
What happens do DNA after death?
- DNA begins to degrade
- Cell death occurs
- Cell membranes rupture & release enzymes that degrade cellular structures
- Chemical reactions to modify & break the DNA strands
- Followed by microbes, both within & external to the body
- DNA degradation occurs over time
DNA begins to degrade
immediately after the cells die
Cell death occurs once a cell is no longer in
contact w/the body’s circulating oxygen supply
Cell membranes rupture & release enzymes that
degrade cellular structures
Chemical reaction act to
modify & break the DNA strands
DNA degradation occurs over time as hydrolytic & oxidative chemical reactions damage the molecules by
fragmentation & chemical modification.
DNA persists in bones & teeth longer than
soft tissues because of rigid structure (provides protection).
DNA binds to
hydroxyapatite of the hard tissues - stabilizes the DNA
Molecular taphonomy is the study of the
various intrinsic & extrinsic factors that influence the degradation of the body’s molecular structures (e.g., DNA).
- Environmental Factors
Differential preservation in different skeletons, in different bones, within the
same skeleton, & even variations in DNA quality & yield across the same bone.
Depositional environment plays a greater role in
degradation than the absolute age.
Bone type & density
contributes to perservation
Different Environmental Factors are
- Temperatures
- Moisture levels
- Oxygen levels
- Microorganism Activity
- Soil Composition
- pH
- Bone Type
Temperature
Autolysis occurs at a maximum chemical activity at
34 degrees C to 40 degrees C
* Two-fold to three-fold increase 10 degree C
Temperature influences microbial activity in that
warmer temp increases activity, thus cooler temps will favor DNA preservation
Some circumstances where mild heating (during extraction) may be beneficial as it makes the
bone brittle, releasing out the DNA.
- Moisture *
Water molecules participate in hydrolytic reactions that
fragment and modify DNA
- Moisture *
The more water/humidity, the more
DNA damage
- Moisture *
Some circumstances where water may influence other environmental variables; for example
peat bog, creates low oxygen environmental and burial in salt water reduce microbial activity.
- Oxygen Levels *
Oxygen molecules participate in oxidative reaction that
modify DNA bases and create lesion in DNA strands.
- Oxygen Levels *
Oxidative reaction leads to degradation as well as
helical distortion (complicates DNA profiling)
- Oxygen Levels *
Oxygen levels influences rate and extent of
microbial decomposition
- Oxygen Levels *
Oxygen rich environments lead to
greater degradation
- Microorgansim Activity*
Microorganism activity can be
endogenous and exogenous
- Microorganism Activity*
Microbes do not digest the DNA directly, instead they digest the protein component of bone, making the DNA
more prone to damage
- Microorganism Activity*
Microbes also produce enzymes that
fragment DNA
- Soil Composition *
Chemical composition complicates
genetic analyzes
- Soil Composition *
Bones/teeth reach chemical equilibrium via mineral leaching and uptake of
different solutes from the soil
- Soil Composition *
Soil composition
degrades bone
- Soil Composition *
Causes chemical changes to be
hydroxyapatite
- Soil Composition *
Soil solutes contain
tannins and humic acids that inhibit PCR
- pH *
Decomp occurs more rapidly in
acidic and alkaline conditions rather than neutral
- pH *
Chemical modifications to hydroxyapatite and DNA influenced by the
pH of the deposition environment
- pH *
Microbial activity also influenced by the
pH of the environment
- Bone Type *
Bone size and construction impact
DNA preservation
- Bone Type *
Larger bones tend to
survive better
- Bone Type *
Dense cortical portions of lower limb bones and harder tissues of teeth are more consistently
reliable in generating DNA profiles as compared to less dense spongy bone.
- Bone Type *
Understanding skeletal elements and those most likely to produce a profile should be considered rather
than sampling based on convenience.
- Biology of Bone *
Rate of degradation of human remains varies greatly with environmental conditions for example
- soft tissue is lost
- Bones are more stable and remain
- Biology of Bone *
Used to identify victims in mass fatalities with techniques such as
- Facial reconstruction
- special body features (scoliosis, implants, replacements, etc.)
- dental records
- DNA analysis
- Biology of Bone *
Adult human skeleton contains
206 bones
- Biology of Bone *
Long Bone shaft is made up of
- outer layer is cortical (compact) bone that is Highly dense, solid & strong
- Middle contains marrow cavity
- Both ends are the epiphysis, consist of cancellous (spongy) bone and is Highly Porous, takes force of compression
- Biology of Bone *
Flat bone is primarily
cortical or cancellous
- Biology of Bone *
Bone contains a
matrix of cells
that makes bone a weight bearing hard tissue
- Biology of Bone *
Inorganic cell matrix consists of
calcium and phosphorus
*hydroxyapatite crystals
- Biology of Bone *
Organic cell matrix consists of
collagens (mostly type I) and are insoluble fibrous proteins
- Biology of Bone *
Developing bones contain
osteoprogenitor cells
- Biology of Bone *
Osteoprogenitor cells produce
osteoblasts
- Biology of Bone *
Osteoblasts produce
new bone matrix
- Biology of Bone *
If osteoblasts are embedded in bone they are
Osteocytes
* most abundant cells in bones
* maintain matrix
* repair damaged bone
- Biology of Bone *
Bone that also contains osteoclasts
dissolve and recycle bone matrix
Large cells that contain 50+ nuclei
- Bone as a Source of DNA *
Most DNA comes from
osteocytes
*Microgram quantities of DNA can be recovered
*Compact bone should contain enough nuclear DNA analysis
- Bone as a Source of DNA *
20,000-26,000 osteocytes
per mm(3) of calcified bone
- Bone as a Source of DNA *
Bones from mass fatalities and burial sites are subjected to
decomposition, nuclear and mtDNA can become degraded
- Bone as a Source of DNA *
Bone sample must be cleaned first by
- Remove contaminating DNA
- Clean outer surface of bone samples prior to processing
- Sanding - used to reduce potential of comingled remains, adhering inhibitors, and bacterial contaminates
- Bone dust from sanding must be cleaned and removed
- Immerse in bleach solution
- Or exposure to UV radiation
- Bone as a Source of DNA *
A high-yield DNA extraction method
- Bone is ground/pulverized into a fine dust for DNA extraction
- .2g-2.5 g bone dust
- Incubate in extraction buffer
- Bone as a Source of DNA *
Incubate in extraction buffer consists of
- Calcified matrix is removed
- Decalcification procedure will dissolve calcium & soften bone tissue
- Proteinase will digest matrix proteins & increase yield.
- Biology of Teeth *
The Two sets of teeth form gestation are
- Deciduous/Primary teeth (20 teeth)
- Secondary/Permanent dentition (32 teeth)
- Bone as a Source of DNA *
Dental records can be used to identify a person by
- Characteristics of teeth
- Alignment
- Overall structure
- X-rays
- DNA from teeth may be used in UHRs
- Bone as a Source of DNA *
Most of a tooth consists of
dentin
- Bone as a Source of DNA *
Crown of a tooth is covered w/
enamel mostly made of collagens