Reduced fitness in interpopulation hybrids can be a first indication of genetic incompatibilities that may ultimately lead to reproductive isolation and speciation. A growing number of cases of hybrid breakdown have been traced to incompatibilities between the nuclear genome and the organellar genomes of the mitochondria and chloroplasts. Although these organellar genomes derive from ancient bacterial endosymbioses, they have been vastly reduced in size and now encode relatively few genes. The remaining genes are necessary but not sufficient for organelle function. In fact, most proteins functioning in the organelles are encoded in the nuclear genome and need to be imported after synthesis in the cytosol. The necessary interactions between organelle and nuclear genomes have resulted in some degree of coadaptation within all natural populations. Hybridization brings together previously untested allelic combinations and can disrupt intergenomic coadaptation, resulting in organelle dysfunction and, consequently, hybrid breakdown.

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As sequencing technologies become more affordable, it is now realistic to propose studying the evolutionary history of virtually any organism on a genomic scale. However, when dealing with non-model organisms it is not always easy to choose the best approach given a specific biological question, a limited budget, and challenging sample material. Furthermore, although recent advances in technology offer unprecedented opportunities for research in non-model organisms, they also demand unprecedented awareness from the researcher regarding the assumptions and limitations of each method. In this review we present an overview of the current sequencing technologies and the methods used in typical high-throughput data analysis pipelines. Subsequently, we contextualize high-throughput DNA sequencing technologies within their applications in non-model organism biology. We include tips regarding managing unconventional sample material, comparative and population genetic approaches that do not require fully assembled genomes, and advice on how to deal with low depth sequencing data.