Sequence analysis of environmental phage isolates.

Stephen C. Hardies, Mandy Rolando, Chris Kawasaki, Borries Demeler, and Philip Serwer. Dept. of Biochemistry, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900.

The high degree of diversity and generally undersampled state of sequences found in bacteriophage genomes is widely acknowledged. Application of large scale genomic sequencing to this problem has thus far been hampered by the nature of phages coming in small genomes, each imposing its own idiosyncratic problems of phage isolation and sequencing of stubborn gaps and ends. We have explored a number approaches to this problem. One is to focus efforts by use of a survey of random shotgun survey of various isolates to select phages of particularly unusual content for full scale sequencing. The complete sequence of phage 1102phi3-1, selected in this way, will be discussed. The properties of this phage as well as others surveyed will be used to explore the issue of how much sequence is needed from a phage to constitute a description of it. Computational approaches will be demonstrated that collapse some of the phage gene diversity into highly diverged phage gene families, thus allowing functional assignment to a higher fraction of sampled genes. In particular, the Sequence Alignment and Modeling System (SAM) (Hughey, Karplus, and Krogh, Technical report at http://www.soe.ucsc.edu/research/compbio/papers/sam_doc/) was found to be particularly adept at aligning highly diverged phage gene subfamilies. An analysis of the large subunit terminase gene family across all tailed phages and Herpes Viruses will be shown as an example of the depth of analysis possible. Finally the problem of establishing phylogenetic relationships among phages will be discussed, both with respect to handling mosaicism and with respect to establishing the absolute time scale for phage gene trees. In particular, the passage of the T7-like RNA polymerase gene into eucaryotes during the mitochondrial endosymbosis will be offered as an ancient event at a known time suitable for establishing rates of evolution. Details with respect to ongoing work in this project can be found at: http://biochem.uthscsa.edu/~hs_lab/phage.html. Supported by grants from the Kleberg Foundation and the Texas Coordinating Board.