TISSUE SAMPLING – HARVESTED MAMMALS (EG. DEER, MOOSE, ELK)
Tissue samples are the ideal sample type for DNA analysis. Tissue samples provide the best quality and quantity of DNA, which helps to facilitate the DNA analyses and provide results in a timely manner. However DNA can be isolated from most biological materials including feces and urine.
Tissue samples located at a kill site are variable. The stability of DNA and quantity is also different within different tissues. The best sample in terms of yield and quality is red muscle. Hide samples with attached tissue are also excellent sources of DNA. DNA in all biological material will degrade until dry, frozen or preserved. When choosing a sample those reddish in colour are best, and this may require burrowing deeper into the tissue in older kill sites.
The DNA in metabolically active tissues such as kidney, spleen, liver, brain and stomach samples undergo rapid enzymatic decay, and should be avoided. Ear clips and closed bones should be taken when located remains are not recent, and the muscle is beginning to decay.
Tissue samples can be stored and shipped at room temperature in the lysis buffer provided in the sample tubes. The lysis buffer dissolves the tissue and inactivates degrading enzymes and prevents microbial activity. The DNA stored this way is stable for up to a year. If no lysis buffer is available, 1 gram (approximately 1 cubic centimeter) of tissue can be packaged in an evidence bag and shipped frozen.
Tissue Sampling Instructions:
- Using CLEAN GLOVES open a new scalpel. Always change gloves between samples to avoid cross contamination.
- If presented with a large piece of meat, burrow into the inside of the meat, to avoid outside contamination (eg. soil). If this is not possible try to shave off a small portion from the outside of the tissue and discard it.
- Then cut a piece of tissue approximately 50-100 mg in mass (approx. 5mm x 5mm x 5mm in dimension) from the centre of the tissue.
- Add the sample to the tube with preservative buffer making sure the entire piece is submerged.
- Ensure that the tube lid is screwed on tight.
- Tissue tubes are often pre-barcoded, and can be used as your item number.
Note: More tissue IS NOT better because there is not enough buffer to digest and preserve a larger piece of tissue. An excessive amount of tissue will decay and DNA integrity will be compromised. Excess tissue samples should be kept frozen at your respective facility as an insurance for potential shipping problems. Multiple samples from an item can be taken and sent for submission. These should be well-labelled as this allows the forensic technician to select the appropriate samples.
Packaging of Tissue
Tissue that has been collected in a tube with lysis buffer can be packaged and shipped at room temperature. To ensure chain of custody, and integrity of your evidence, place your tissue samples into an evidence bag and record the required details. Place evidence from a suspect into separate evidence packages to your evidence collected from a kill site. This is to help ensure that there is no contamination between the samples you would like compared. Large pieces of meat have been known to thaw and leak through evidence bags, which can compromise surrounding evidence.
TISSUE SAMPLING – FISH
Fish blood contains nucleated red blood cells and small amounts of fish blood provide larger amounts of DNA than mammalian blood. Investigations involving fish can involve the discovery of processed gut piles, cleaned individual fish, and fillets with no distinguishing features. As with investigations involving mammals, tissue samples provide good quality and quantity of DNA.
In the case of whole fish, fin clips (pectoral, pelvic, adipose or caudal) are good samples to collect as they contain capillaries with good amounts of blood. As with mammal samples internal tissues should be avoided.
Fillet tissues have more water content than red muscle tissue. As such, approximately 100-200mg of tissue should be collected and submitted in a tissue tube following the same tissue collection instructions as above. In determining the number of fish seized in the discovery of fillets, a sample from each fillet should be collected.
Fish tissue samples should be sent in tissue tubes whenever possible. A small amount of tissue or individual fillets (depending on size) can be packaged in evidence bags or sealed Ziploc containers (depending on number of fillets) and shipped frozen with ice packs if tissue tubes are not available.
When sampling hair, it is important that hair is plucked and not cut, so that the root of the hair shaft is collected. Without a root, species identification using mitochondrial DNA is often the only DNA analysis that can be performed (hair shaft only contains mitochondrial DNA). The root of the hair contains nuclear DNA, which is required to test for gender and individual identification. The ideal hair samples are found on the back of the shoulder where the hair is thick and has large roots. 15-20 hairs with roots should be collected to provide enough DNA for obtaining a full individual profile.
Hair samples should be kept dry and shipped in envelopes at room temperature. A desiccant pouch should be placed in the evidence bag or Ziploc with the envelope. The desiccant pouch prevents moisture from degrading the DNA.
Hair Sampling Instructions
- Using CLEAN GLOVES, open a new pair of disposable forceps and collect the hair evidence.
- If you have a hair envelope, place the hairs on the double sided tape, making sure not to stick the roots onto the tape. If you do not have a hair envelope, a coin envelope can be used. Plastic should be avoided as it retains moisture and degrades the DNA.
- Label the necessary details onto the envelope and place it into an evidence bag.
- Place a desiccant pouch into the evidence bag to reduce moisture.
These instructions also apply to feathers. Plucked feathers should be recovered first before any shed feathers, as these have more DNA at the root. Any blood at the end of feathers is also a good source of DNA. Like fish, amphibians and reptiles; bird blood contains nucleated blood cells so a small amount gives a high yield.
Blood is a good source of DNA. Blood samples located on materials such as clothing, knives, guns, arrows etc. should be shipped at room temperature in a well sealed package. Keep in mind that blood stains may have to be cut or isolated from the material submitted. If blood stains are located on an object that cannot be shipped (eg. On an ATV), follow these steps:
Blood Sampling Instructions
- Using CLEAN GLOVES moisten a swab with Phosphate Buffered Saline (PBS). If this is not available, distilled water can be used.
- Collect the stain and air dry the swab before placing it back into the tube.
- Place the swab into an evidence bag and record the necessary details.
- Swabs should be shipped dry and at room temperature.
Blood stains from a kill site are commonly found on leaves or rocks. If the stain is large enough, sample the blood using a dry swab without contaminating it with leaf material or dirt. Air-dry the swab for a few minutes, and place it into the sealed tube provided and ship at room temperature.
Small blood stains located on items such as twigs or leaves can be placed into a 50mL desiccant tube. Label the tube with your item number and place it into an evidence bag and ship at room temperature.
Blood located in frozen snow and ice should be collected in a vial/tube. Keep the sample frozen and ship the sample with ice packs. Approximately 5-10 milliliters of the snow should be sent.
TEETH, BONE AND HORN SAMPLING
Samples of teeth, bone and antlers can be used for DNA extraction and analysis. When fresh or recent tissue from a carcass is unavailable, long bones, teeth or antlers are a better sample choice (along with ear clips). Bones and teeth without any tissue attached can be placed into evidence bags and shipped at room temperature. If there is any tissue connected, ship the item frozen/cold with ice packs.
Antler shavings or a portion of antler can be packaged and submitted at room temperature. Follow these steps if you submit antler shavings:
- Using CLEAN GLOVES use a sterilised drill bit to remove antler shavings.
- Drill and collect shavings from the interior as best as possible. This is to avoid contamination from the outer surface.
- Place the shavings into a sealed container, such as a pill bottle.
- Seal the container and label the item. Ship at room temperature.