Biological Sciences, School of


Date of this Version

Fall 12-4-2009

Document Type



A DISSERTATION Presented to the Faculty of The Graduate College at the University of Nebraska In partial Fulfillment of Requirements For the Degree of Doctor of Philosophy, Major: Biological Sciences; Under the Supervision of Professor James L. Van Etten
Lincoln, Nebraska: December, 2009
Copyright (c) 2009 Giane M. Yanai


Paramecium bursaria chlorella virus (PBCV-1), a member of the family Phycodnaviridae, is a large dsDNA, plaque-forming virus that infects the unicellular green alga Chlorella NC64A. The 331 kb PBCV-1 genome is predicted to encode 365 proteins and 11 tRNAs. To follow global transcription during PBCV-1 replication, a microarray containing 50-mer probes to the PBCV-1 365 protein-encoding genes (CDS) was constructed. Competitive hybridization experiments were conducted employing cDNAs from poly A-containing RNAs obtained from cells at seven time points after virus infection. The results led to the following conclusions: i) the PBCV-1 replication cycle is temporally programmed and regulated; ii) 360 (99%) of the arrayed PBCV-1 CDSs were expressed at some time in the virus life cycle in the laboratory; iii) 227 (62%) of the CDSs were expressed before virus DNA synthesis begins; iv) these 227 CDSs were grouped into two classes: 127 transcripts disappeared prior to initiation of virus DNA synthesis (considered early) and 100 transcripts were still detected after virus DNA synthesis begins (considered early/late); v) 133 (36%) of the CDSs were expressed after virus DNA synthesis begins (considered late); vi) expression of most late CDSs is inhibited by adding the DNA replication inhibitor, aphidicolin, prior to virus infection. This study provides the first comprehensive evaluation of virus gene expression during the PBCV-1 lifecycle.

Adviser: James Van Etten