Sampling of tooth roots for ancient DNA
Marcel Keller, Christiana L Scheib
Abstract
Protocol for the sampling of tooth roots of archaeological human remains for the extraction of ancient DNA.
Before start
Previous step:
This protocol follows the introduction of samples into the ancient DNA lab (documentation and changing bags).
Following step:
Proceed with the decontamination protocol.
Equipment and consumables:
| A | B |
|---|---|
| Number | Equipment and consumables |
| 4 | disposable 100 ml beakers |
| 2x100 ml | NaOCl (6% v/v) |
| 100 ml | MilliQ water |
| 100 ml | 70% ethanol |
| 2 of each | drill disks, tweezers, dental scalers, tweezers |
| 1 | vice |
| 1 | toothbrush |
Steps
Preparation
Turn the hood on full power and open the glass.
Clean drill hood and table bench surfaces with DNA Exitus and wipe clean with a paper towel.
Set up decontamination station:
4 disposable 100 ml beakers
6% NaOCl (bleach) aliquot [2x]
MilliQ water [1x]
70% ethanol aliquot [1x]
To decontaminate drill bits, drill spanner, tweezers, dental scalers and toothbrush – Soak in 6% bleach for 5 minutes, rinse with MilliQ water, rinse with 70% Ethanol. Clean the drill spanner and tooth extractors with DNA Exitus.
Wipe down markers and pens with DNA Exitus.
Label your bags and tubes:
| A | B |
|---|---|
| Bags for drilled samples | |
| Sample ID | ABC001A |
| Date | DD/MM/YYYY |
| Initials | YZ |
| A | B | C |
|---|---|---|
| Tubes e.g. for collecting calculus | Top | Side |
| Content | e.g. calculus/drill powder | |
| Project ID | PROJ | PROJ |
| Sample ID | ABC001A | ABC001A |
| Date | DD/MM/YYYY | DD/MM/YYYY |
| Initials | YZ | YZ |
Place down sheets of aluminum foil to cover the entire surface of the drill hood.
Make an L-shaped “wall” of folded foil that runs along the back of your work area and the side without the drill to keep flying dust from expanding across the hood.
Place down a smaller piece of foil for your working surface. Fold small pieces of foil that you can wrap around the beaks of the tooth extractor. Place a big weighing boat onto the working surface. Change this setup for each sample.
Gather clean, decontaminated tools for drilling and place them on the foil.
Sampling
Turn on the drill in “FWD” mode and choose a medium speed to prevent overheating/burning of the bone or dentine.
Put on clean sleeves by pushing your thumbs through the plastic close to the cuffs to keep the sleeve in place, then gloves on top.
Place the bag with your sample and the clean sample bag and tube into the hood behind the foil wall.
If the tooth has calculus, use the dental scaler to remove it before sampling. Put the tooth into the mini grip bag and hold the tooth outside the bag with your fingers. Now push gently with scaler and collect all calculus inside the bag. Remove the tooth and pour the calculus on weighing paper. Use weighing paper to place calculus in a labeled 1.5 ml tube after taring.
Wrap foil strips around the ends of your tooth extractor (for holding the tooth).
Hold the tooth with the tooth extractor and drill carefully a few millimeters above the end of the root.
When drilling is completed, put your samples in labeled bags. Wipe these down with a small amount of DNA Exitus before adding to your sample box.
Clean-up after each sample
Rinse off the drill wheel with MilliQ water first then into your 6% bleach for a few minutes or while you are drilling the next tooth. Then rinse with water and finally ethanol before drying for use again.
Carefully remove and throw away the weighing boat, small work surface foil, dirty gloves, and sleeves. Put on new gloves.
For the next sample, start from step 9.
Final clean-up
When finished for the day, wipe down the tooth extractors and drill spanner with DNA Exitus and put away.
Use a clean toothbrush with a little DNA Exitus to remove sample powder from the top/inside of the drill head.
Leave drill bits and scalers to soak in bleach while you throw away all foil and other consumables and wipe down all inner hood work surfaces and the outside of the drill with DNA Exitus and wipe dry with paper towels.
