DArT (Diversity Arrays Technology)
DArT is one of the recently developed molecular techniques and it has only been used in rice, barley, eucalyptus, Arabidopsis, cassava, wheat and pigeonpea. The inventors promote it as an open source (non- exclusive) technology with a great potential for genetic diversity and mapping studies in a number of 'orphan' crops relevant in Third World countries. DArT is a microarray hybridization-based technique that enables the simultaneous typing of several hundred polymorphic loci spread over the genome. Details of the methodology for DArT was first described by Jaccoud et al. (2001). For each individual DNA sample being typed, genomic representations are prepared by restriction enzyme (e.g., PstI and TaqI) digestion of genomic DNA followed by ligation of restriction fragments to adapters. The genome complexity is then reduced by PCR using primers with complementary sequences to the adapter and selective overhangs. The fragments from representations are cloned, and cloned inserts are amplified using vector-specific primers, purified and arrayed onto a solid support (microarray) resulting in a "discovery array." Labeled genomic representations prepared from the individual genomes included in the pool are hybridized to the discovery array (Jaccoud et al., 2001). Polymorphic clones (DArT markers) show variable hybridization signal intensities for different individuals. These clones are subsequently assembled into a "genotyping array" for routine genotyping.
DArT technique has a number of advantages:
a. It does not need prior sequence information for the species to be studied; this makes the method applicable to all species regardless of how much DNA sequence information is available for that species.
b. It is a high throughput, quick, and highly reproducible method.
c. It is cost effective, with an estimated cost per data point tenfold lower than SSR markers (Xia et al., 2005).
d. The genetic scope of analysis is defined by the user and easily expandable.
e. It is not covered by exclusive patent rights, but on the contrary open-source (i.e., it is designed for open use and shared improvement).
This technique, however, has also its own limitations:
1. DArT is a microarray-based technique that involves several steps, including preparation of genomic representation for the target species, cloning, and data management and analysis. The latter requires dedicated software's such as DArTsoft and DArTdb. The establishment of DArT system, therefore, is highly likely to demand an extensive investment both in laboratory facility and skilled manpower.
2. DArT assays for the presence (or amount) of a specific DNA fragment in a representation. Hence, DArT markers are primarily dominant (present or absent) or differences in intensity, which limits its value in some applications.
3. The technology has been used in few species primarily by the team that developed it (who has setup a quite economical commercial service for some species); only a single independently group has so far successfully established the methodology to Eucalyptus grandis in South Africa (Lezar et al., 2004).
The Purpose Marker
In the previous sections, we have reviewed the principle and methodology of 11 different types of molecular markers that have been used for different purposes in plants. The main challenges for researchers would, therefore, be selecting one or more of these markers for their specific project. The desirable properties of molecular markers are high polymorphism, codominant inheritance, frequent occurrence and even distribution throughout the genome, selectively neutral behavior, easy access, easy and fast assay, low cost and high throughput, high reproducibility, and transferability between laboratories, populations and/or species. No molecular markers are available yet that fulfill all these requirements. However, according to the kind of study to be undertaken, one can choose among the variety of molecular marker systems, each of which combines at least some of these desirable properties. A number of factors need to be considered in choosing one or more of the various molecular marker types:
a) Marker system availability.
b) Simplicity of the technique and time availability.
c) Anticipated level of polymorphism in the population.
d) Quantity and quality of DNA available.
e) Transferability between laboratories, populations, pedigrees and species.
f) The size and structure of the population to be studied.
g) Availability of adequate skills and equipment.
h) Cost per data-point and availability of sufficient funding.
i) Marker inheritance (dominant versus codominant) and the type of genetic information sought in the population
DArT is a recent technique and it remains to be thoroughly tested in various species.
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