Unknown,Transcriptomics,Genomics,Proteomics

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Cell cycle-regulated gene expression in tobacco cells

ABSTRACT: Tobacco BY-2 cells can be highly synchronized by treatment with an inhibitor of DNA synthesis, aphidicolin. For comprehensive understanding of cell cycle-regulated gene expression in tobacco, microarray analysis was conducted in synchronized cultures of BY-2 cells. Cells were treated with aphidicolin for 24 hours to block the entry into and progression through the S phase. After release from the aphidicolin block, cells were sampled every 2 h from 0â16 h. cDNA probes prepared from these samples were hybridised to a custom-made16K microarray. We performed this procedure twice independently to identify transcripts that reproducibly showed similar cell cycle-regulated expression. A total of 883 putative cell cycle-regulated transcripts were identified from 16,896 duplicated spots on the microarrays. These transcripts could be categorised into 10 classes on the basis of transcript expression patterns in the cell cycle. Keywords: time course Synchronization experiments were repeated twice (experiments A and B). In each experiment, cells were sampled every 2 h during the culture periods (0-16 h), and used for microarray analysis.

ORGANISM(S): Nicotiana tabacum

SUBMITTER: Hiroo Fukuda

PROVIDER: E-GEOD-14250 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description: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 cDNA from poly A-containing RNA 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 are expressed at some time in the virus life cycle in the laboratory; iii) 227 (62%) of the CDSs are expressed before virus DNA synthesis begins; iv) these 227 CDSs were grouped into two classes: 127 transcripts disappear prior to initiation of virus DNA synthesis (considered early) and 100 transcripts are still detected after virus DNA synthesis begins (considered early/late); v) 133 (36%) of the CDSs are 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. Time course analysis of chlorella virus PBCV-1: cDNAs from poly A-containing RNAs isolated from cells at seven times after PBCV-1 infection (20, 40, 60, 90, 120, 240, 360 min p.i.) were competitively hybridized against a reference sample on the microarrays. Three independent biological replica, four technical replica of the genome per array. Time course analysis of chlorella virus PBCV-1 gene expression in the presence of aphidicolin: cDNAs from poly A-containing RNAs isolated from aphidicolin-treated cells at seven times after PBCV-1 infection (20, 40, 60, 90, 120, 240, 360 min p.i.) were competitively hybridized against non-treated, infected samples from the same time points. Three independent biological replica, four technical replica of the genome per array.

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Cell cycle-regulated gene expression in tobacco cells

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