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Background  Microalgae are promising feedstock for production of lipids, sugars, bioactive compounds and in particular biofuels, yet development of sensitive and reliable phylotyping strategies for microalgae has been hindered by the paucity of phylogenetically closely-related finished genomes.    Results  Using the oleaginous eustigmatophyte  Nannochloropsis  as a model, we assessed current intragenus phylotyping strategies by producing the complete plastid (pt) and mitochondrial (mt) genomes of seven strains from six  Nannochloropsis  species. Genes on the pt and mt genomes have been highly conserved in content, size and order, strongly negatively selected and evolving at a rate 33% and 66% of nuclear genomes respectively. Pt genome diversification was driven by asymmetric evolution of two inverted repeats (IRa and IRb):  psbV  and  clpC  in IRb are highly conserved whereas their counterparts in IRa exhibit three lineage-associated types of structural polymorphism via duplication or disruption of whole or partial genes. In the mt genomes, however, a single evolution hotspot varies in copy-number of a 3.5 Kb-long,  cox1 -harboring repeat. The organelle markers (e.g.,  cox1 ,  cox2 ,  psbA ,  rbcL  and  rrn16_mt ) and nuclear markers (e.g.,  ITS2  and  18S ) that are widely used for phylogenetic analysis obtained a divergent phylogeny for the seven strains, largely due to low SNP density. A new strategy for intragenus phylotyping of microalgae was thus proposed that includes  (i)  twelve sequence markers that are of higher sensitivity than  ITS2  for interspecies phylogenetic analysis,  (ii)  multi-locus sequence typing based on  rps11_mt - nad4 ,  rps3_mt  and  cox2 - rrn16_mt  for intraspecies phylogenetic reconstruction and  (iii)  several SSR loci for identification of strains within a given species.    Conclusion  This first comprehensive dataset of organelle genomes for a microalgal genus enabled exhaustive assessment and searches of all candidate phylogenetic markers on the organelle genomes. A new strategy for intragenus phylotyping of microalgae was proposed which might be generally applicable to other microalgal genera and should serve as a valuable tool in the expanding algal biotechnology industry.

Nannochloropsis plastid and mitochondrial phylogenomes reveal organelle diversification mechanism and intragenus phylotyping strategy in microalgae