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10. MARKER EFFICIENCY
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obvious applications taken into account the different technological input necessary for the
development of each marker category. The usefulness of each marker category is presented
in the three important areas of genetic research - these areas which were also touched in the
studies on Lolium evolution.
10.2. COSTS OF MARKER ASSAYS
When starting any analysis, everyone would like to obtain as much information as possible
at as low costs and personal efforts as possible. Hence, it is not surprising that the first
thing to consider is the number of markers (bands) that should be generated and techniques
that are the cheapest with respect to input and information retrieved. Despite chemicals
needed for a particular reaction, matrices through which small DNA or protein fragments
migrate should be considered as well. The usual electrophoretic media are agarose and polyacrylamide
gels for both DNA and proteins in addition to starch for the latter. From a technical
point of view, any marker system that requires polyacrylamide gel electrophoresis is more
challenging that those based on agarose or starch gel. On the other hand, polyacrylamide
gives the highest resolution, but not always it is necessary. Notwithstanding these methodological
considerations, another popular opinion is that enzymes are the cheapest while DNA
markers such as AFLP or transposon-based (SSAP) belong to the most expensive. It is true
if total costs per a sample are taken into account (Figure 10.1). Roughly, a single AFLP or
SSAP reaction together with costs of a gel and staining is almost sevenfold more expensive
than enzyme or DNA analyses such as RAPD. However, the proportion becomes inverted if
the total number of revealed loci is considered. To obtain 100 bands or loci one should pay
about 10 euros if AFLP or SSAP markers are used but 100 enzymatic loci would cost about
40 euros if it was possible to obtain (Figure 10.2). Indeed, even RAPD markers are more
expensive then AFLP/SSAP. The analysis of cpDNA and mtDNA, so popular in phylogenetic
studies, also is not cost effective especially when coupled with restriction digestion. These
differences are even more dramatic if polymorphic loci are taken into account (Figure 10.3).
It is obvious from these comparisons that such techniques as AFLP or SSAP are the most
cost effective. Another advantage is that the huge number of markers can be obtained with
less man power. A single AFLP or SSAP gel contains from 50 to 100 bands, so only a few
analyses are needed to sample a large portion of loci. For example, a single RAPD primer
can generate up to seven loci, among which about six can be polymorphic (Figure 10.4). In
comparison, a pair of SSAP primers can give from 60 to 90 bands and about 40 to 80 are
polymorphic (Figure 10.5). So it is feasible to assay hundreds of individuals for a hundred of
polymorphic loci in a week or so without much effort. On the other hand, they need a polyacrylamide
gel and the start-up costs are quite high. In a case the AFLP or SSAP are too demanding
and agarose gel should be used instead, the markers based on junctions between
introns and exons (ISJ) are recommended due to relatively high polymorphism. Nonetheless,
RAPDs are more cost effective that cpDNA or enzymes. To this point the possibility to obtain
enough DNA may be a certain limitation (Figure 10.6). Although amount of DNA per a reaction
is exactly the same for RAPD and ISJ, the latter markers are more polymorphic and
hence, total amount of DNA to obtain 100 polymorphic loci is lower.