1. www.le.ac.uk/.../ar99-00/international/ international.html
Applications of Molecular Biology in Archaeology - Extraction of Ancient DNA Bradwell, S. and Nguyen, R.
international.html

2. BACKGROUND What is Ancient DNA?
Ancient DNA refers to DNA obtained from highly degraded ancient biological remains (such as bone, teeth and hair).
For a long time, scientists did not expect long-term DNA-preservation in ancient remains
because degradation results shortly after individual’s death.

3. BACKGROUND
Environmental Influences

4. BACKGROUND
Early 1980’s
Investigated mummies as old as 2,400 yr-old for DNA and found that DNA could be molecularly cloned in a plasmid vector.
Problem:
Classical molecular biological methodologies required high amounts of DNA material, but most ancient DNA are highly degraded and only minute quantities may be extracted
news.nationalgeographic.com/.../ 07/0709_mummycongress.html

5. Polymerase-Chain Reactions
Field of Molecular Archaeology was “revolutionized” by the invention of PCR
Technique made it possible to detect and characterized minute traces of DNA.
Ancient-DNA research has been a proven to be a powerful tool used in archaeological study.
mtdna.html

6. Extraction
Teeth and bones are excellent sources for the extraction of ancient DNA.

7. Extraction Begin by breaking up tissue 1) Grinding in liquid nitrogen
2) Classic phenol-chloroform extraction
3) Salt-based extraction
4) A variety of commercial kits
EN/ html
mar/tools_ html
forensic.to/webhome/dna/paternity.htm

8. PCR Success
Successful DNA typing of both short tandem repeats loci and the hypervariable region of the human mitochondrial DNA was achieved after extracting total DNA from bones
Two common methodologies employed:
1) Centrificon approach
2) QIAquick approach

9. www.pharmcentral.com/ nucleotides.htm
Problems with PCR
More than 99% of ancient DNA are chemically modified
SOURCES OF DAMAGE:
Hydrolytic damage resulting in deamination of bases
2) Oxidative damage
3) Presence of unidentified inhibitors of PCR
nucleotides.htm

10. www.cnn.com/2000/NATURE/11/ 10/ancient.europeans.ap/
Solutions
Knowledge! - The size of the amplification product inversely correlates with the amplification efficiency
Gas chromatography and Mass Spectrometry allow the determination of a damage threshold
Verify DNA authenticity through the estimation of D/L ratio of amino acids
Simulate environmental conditions
10/ancient.europeans.ap/

11. Risk of contamination with contemporary DNA
More Problems with PCR
Risk of contamination with contemporary DNA
Allelic drop out
index.asp?mpage=iqualicon&c=UK

12. www.aero.org/facilities/ environmentalApps.html
Solutions
Common sense
Sample pre-treatment
Use of controls
environmentalApps.html

13. Case Study
The Discovery of the Kwäday Dän Ts’ìnchi Ancient Remains at the Edge of a Glacier in British Columbia’s Tatshenshini-Alsek Park.

14. Kwäday Dän Ts’ìnchi = person long ago found
Case Study
Kwäday Dän Ts’ìnchi = person long ago found
“Our oral history has told us about the importance of this area as a travel route and about the nearby villages on the Tatshenshini River. Now Kwaday Dän Sinchì brings these stories to life.”

15. Case Study
Initial radiocarbon dating indicates that the remains are approximately 550 years old

16. Case Study
First access was given to the researchers Monsalve et al. 2002
DNA extracted from the humerus, overlaying muscle, bone collagen
Proteinaceous material was detected by measuring the % of C and N in whole bone and collagen-type residue using an elemental analyzer
Denaturation step of PCR cycle lengthened to 5 min

17. Case Study Ancient DNA authenticity determined by:
1) Verifying nitrogen and carbon % in hard and soft tissue met requirements for preservation of proteinaceous material
2) Analyzing mtDNA sequences from PCR fragments and cloning products of the hypervariable control region
3) Validating the humerus DNA results with an independent study

18. Conclusion Ancient DNA provides insight into historical events
It has been successfully applied for understanding ancient pathologies
Finding may also be used for the reconstruction of population origins
Ancient DNA has been applied to determine evolutionary relationships as well as paternity and maternity

19. The End