Mitochondrial nucleotide sequences, in contrast to their plastid counterparts, have been rarely exploited for use in determining algal phylogenetic relationships. Their utilization would prove advantageous particularly for the assessment of evolutionary relationships among protistian lineages that include photosynthetic species, their colorless heterotrophic descendents which contain only a reduced plastid genome, as well as ancestral non-photosynthetic relatives. Recent studies that have examined mitochondrial protein (cox1, cox2, cob, and nad5), rather than ribosomal subunit, genes confirm that these algal sequences are sufficiently conserved to be used routinely in algal systematics. In this investigation we sequenced the mitochondrial nad4L gene, encoding a small (ca. 99 amino acid) polypeptide subunit of the NADH dehydrogenase complex, from two chromophytic algae and an oomycete to determine its suitability for use in determining algal and protistan evolutionary relationships. Our analysis demonstrated that nad4L could clearly resolve these three organisms as a distinct clade, the stramenopiles, from terrestrial plants/chlorophytes and eufungi. However, the position of deeply rooted species within the stramenopiles proved tenuous and would best be examined in conjunction with the analysis of longer mitochondrial protein gene sequences.