PCR is the first step in most genetic analyses, from genetic fingerprinting to creat-
ing genetic phylogenies.
For studies of phylogeny, highly conserved mitochondrial DNA is used (Parker
et al. 1998). For identification of individuals it is the highly variable DNA found in
the major histocompatibility complex(MHC) that is used. Once a target region has
been chosen, a short piece of DNA is synthesized to act as a primer (a piece onto
which new DNA is attached). We start with a mixture of the original double-
stranded DNA, the primers, free nucleotides, and a heat-stable DNA polymerase. The
mixture is heated and the strands of the double DNA are separated. Upon cooling,
the primers attach themselves at one end of the target DNA and serve as starting
points from which the polymerase builds the copy. A new cycle of heating starts the
process again and is repeated for 25 – 49 rounds, depending on the protocol, to make
over a million copies of the selected DNA region.
PCR has been used to amplify DNA from extinct animals so as to elucidate phy-
logenetic relationships. Samples of ancient DNA from extinct species show how the
quagga (Equus quagga) is placed in the zebras (Higuchi et al. 1984) and the saber-
tooth cat (Smilodon) is within the Felidae ( Janczewski et al. 1992). Samples from the
moas (extinct ratites of New Zealand) show that they are an ancient lineage not closely
related to modern-day kiwis (Apteryx). This indicates that there were two invasions
of New Zealand by ratites (Landweber 1999). PCR was also used to show that genetic
variation in humpback whales (Megaptera novaeangliae) was not reduced when the
population went through low numbers from commercial harvesting (Baker et al. 1993).
PCR has replaced several older techniques that are now going out of use. For
historical interest some of these are restriction fragment length polymorphisms
(RFLPs), random amplified polymorphic DNA (RAPDs), and variable number
tandem repeats(VNTRs). RFLPs take advantage of mutations in the DNA that can
be detected by the presence or absence of a cleavage site revealed by an enzyme called
a restriction endonuclease. Restriction enzymes cleave or cut the DNA at particular
recognition sites located at random along the DNA molecule. Total DNA is isolated
from a tissue sample, challenged with restriction enzymes, and then electrophoresed
on an agarose gel matrix. Differences between individuals are detected from the
distribution of fragment lengths. This technique was used to detect genetic disorders
in humans such as thalassemia (Weatherall 1985), and in studies of the reproduc-
tive behavior of lesser snow geese (Cheu caerulescens) in northern Canada (Quinn
et al. 1987).
The use of RAPDs for population genetic inference is being questioned because of
problems with reproducibility, dominance, and homology. They can be used for genetic
mapping studies or for species diagnostic markers.
Mitochondrial DNA techniques
Mitochondria in cells have their own DNA (mtDNA) whose strands are relatively short
(1.6 × 104 base pairs compared with 10^9 base pairs for the nuclear DNA). Parts of
the mtDNA mutate at a fast rate and are highly variable, such as the MHC or the d-
loop (though tandem repeat areas of nuclear DNA are even more variable). Regions of
mtDNA can be monitored for mutations with radioactive probes in the same way as
nuclear DNA. Genetic variability accumulates rapidly and large differences between
populations are thereby often evident. mtDNA is inherited by matrilineal descent only,
thus permitting an assessment of novel sources of variability. For example, there are