A MITE-based genotyping method to reveal hundreds of DNA polymorphisms in an animal genome after a few generations of artificial selection
1 Laboratoire d'Ecologie Alpine, CNRS-UMR 5553, Université Joseph Fourier, BP 53, 38041 Grenoble cedex 09, France
2 Diversity Arrays Technology P/L, PO Box 7141, Yarralumla, ACT 2600, Australia
BMC Genomics 2008, 9:459 doi:10.1186/1471-2164-9-459Published: 6 October 2008
For most organisms, developing hundreds of genetic markers spanning the whole genome still requires excessive if not unrealistic efforts. In this context, there is an obvious need for methodologies allowing the low-cost, fast and high-throughput genotyping of virtually any species, such as the Diversity Arrays Technology (DArT). One of the crucial steps of the DArT technique is the genome complexity reduction, which allows obtaining a genomic representation characteristic of the studied DNA sample and necessary for subsequent genotyping. In this article, using the mosquito Aedes aegypti as a study model, we describe a new genome complexity reduction method taking advantage of the abundance of miniature inverted repeat transposable elements (MITEs) in the genome of this species.
Ae. aegypti genomic representations were produced following a two-step procedure: (1) restriction digestion of the genomic DNA and simultaneous ligation of a specific adaptor to compatible ends, and (2) amplification of restriction fragments containing a particular MITE element called Pony using two primers, one annealing to the adaptor sequence and one annealing to a conserved sequence motif of the Pony element. Using this protocol, we constructed a library comprising more than 6,000 DArT clones, of which at least 5.70% were highly reliable polymorphic markers for two closely related mosquito strains separated by only a few generations of artificial selection. Within this dataset, linkage disequilibrium was low, and marker redundancy was evaluated at 2.86% only. Most of the detected genetic variability was observed between the two studied mosquito strains, but individuals of the same strain could still be clearly distinguished.
The new complexity reduction method was particularly efficient to reveal genetic polymorphisms in Ae. egypti. Overall, our results testify of the flexibility of the DArT genotyping technique and open new prospects as regards its application to a wider range of species, including animals which have been refractory to it so far. DArT has also a role to play in the current burst of whole-genome scans carried out in various organisms, which track signatures of selection in order to unravel the basis of genetic adaptation.