Biol. Bull. 207: 169. (October 2004)
© 2004 Marine Biological Laboratory
Initial Sequence and Protein Modeling Results of a Mitochondrial Genome Project on Understudied Invertebrate Phyla
Gillian E. Robbins1,2,
Gonzalo Giribet3,
Karin Kiontke4,
David H. Fitch4,
Jeffrey L. Boore5 and
Robert K. Campbell1,2
1 Serono Reproductive Biology Institute, Rockland, Massachusetts
2 Marine Biological Laboratory, Woods Hole, Massachusetts
3 Harvard University, Cambridge, Massachusetts
4 New York University, New York, New York
5 DOE Joint Genome Institute, Walnut Creek, California
Mitochondria play an essential role in metabolism. Comprising its own circular DNA, the mitochondrial genome of metazoans typically encodes 2 rRNAs, 22 tRNAs, and 12 to 13 polypeptides. These polypeptides are responsible for the production of protein complexes in the inner mitochondrial membrane that facilitate the production of ATP. Based upon genomic similarities, eukaryotic mitochondria may have evolved from an alphaproteobacterium-like ancestor.
A mitochondrial genomics study was initiated using organisms from invertebrate phyla including Priapulida (Halicryptus spinulosus), Rotifera (Encentrum tectipes), Acanthocephala (Echinorynchus gadi), Gnathostomulida (Gnathostomula peregrini), Cycliophora (Symbion pandora), and Nematoda (Oscheius tipulae and Rhabditoides regina). Gene regions from cytochrome c oxidase I (coxI), cytochrome b, 12S rRNA and 16S rRNA were amplified and sequenced using universal primer pairs. Nested primers were then designed to amplify larger fragments of each mitochondrial genome. Nested PCR products (5–10 kb) were obtained from three organisms, which covered 50%–75% of the genomic mtDNA in highly purified form. These products will be subjected to shotgun sequencing. Further PCR studies of the remaining fragments, as well as those of other mitochondrial genomes, are ongoing.
Successful amplification of coxI sequences using LCO/HCO primers was achieved from all invertebrates studied. Cytochrome c oxidase, the terminal oxidase in the respiratory chain, reduces oxygen to water through electron and proton transfer. The PCR of the coxI from the rotifer (E. tectipes) sample yielded two divergent protein-coding sequences. BLAST analyses revealed one to be most closely related to other rotifer coxI sequences. The second sequence appears equally related to coxI genes from alphaproteobacteria (e.g., Rickettsia) and from various eukaryotes, suggesting that this gene and its associated genome could be in an interesting phylogenetic position.
