Biology Reference
In-Depth Information
Theoretical calculations and empirical observations suggest DNA should only
be able to survive, in a highly fragmented and chemically modified form, for
50,000-100,000 years (
Austin et al. 1997a, Rollo 1998, Hofreiter et al. 2001
).
Because only tiny amounts of DNA usually can be extracted from an archeologi-
cal specimen, stringent precautions and multiple controls are required to avoid
accidental contamination with modern DNA.
A methodology to deal with ancient specimens has been proposed that
includes careful selection of well-preserved specimens, choice of tissue samples
that are likely to have the best DNA preservation, and surface sterilization to
eliminate surface contamination (
Hebsgaard et al. 2005
). The operations should
be carried out in a laboratory dedicated to work on ancient specimens and work
on ancient DNA should be separated from that on modern DNA (
Austin et al.
1997a, Cooper and Poinar 2000, Hofreiter et al. 2001
). Most importantly,
mul-
tiple
negative controls should be performed during DNA extraction and PCR set
up, although a lack of amplifications in the negative controls is not definitive
proof of authentic ancient DNA.
Another consideration is the likelihood of amplifying nuclear copies of
mitochondrial genes (
den Tex et al. 2010
). Universal primers are often used to
amplify DNA from mitochondrial genes in ancient specimens. However,
den Tex
et al. (2010)
found that nuclear copies were often amplified instead of mito-
chondrial DNA, especially when universal primers were used.
Another crucial step is the authentication of the results. Putatively ancient DNA
sequences should be obtained from different extractions of the same sample
and from different tissue samples from different specimens (
Austin et al. 1997a,
Cooper and Poinar 2000
). The ultimate test of authenticity should be independent
replication in two separate laboratories (
Rollo 1998
). So far, this type of replica-
tion has not been achieved for DNA from amber-preserved arthropod specimens
(
Austin et al. 1997b, Walden and Robertson 1997, Gutierrez and Marin 1998
).
Thomsen et al. (2009)
report a method for nondestructive sampling of ancient
insect DNA, which allowed insects from a museum collected in 1820 to be ampli-
fied, but was less successful with samples from permafrost sediments that were
3280-1800 years old.
8.5.2 Amplifying Old DNA
Amplification of old DNA from museum specimens is less difficult and less con-
troversial (
Paabo 1990, 1991; Jackson et al. 1991; Cano et al. 1993a,b; Townson
et al. 1999; Watts et al. 2007
). DNA from pathogens contained within museum
specimens of arthropods can be amplified by the PCR. For example, Lyme
