DNA replication chronology and replicon mapping of developmental genes in Physarum polycephalum /
| Main Author: | |
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| Other Authors: | , , |
| Format: | Thesis Book |
| Language: | English |
| Published: |
1991.
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| Subjects: | |
| Online Access: | Link to OAKTrust copy |
| Abstract: | Transcriptional activity has been correlated with the early DNA replication of genes during the S phase of the cell cycle in eukaryotes. Early-replicated genes are typically active while genes replicated later tend to remain silent. This correlation has been extended using cDNA sequences of developmentally regulated genes from different life cycle stages of the myxomycete Physarum polycephalum in replication timing assays. Timing assays were developed using isolated nuclei, derived from multinucleate Physarum plasmodia, which traverse the cell cycle synchronously. Synchronous DNA samples were also analyzed by two-dimensional (2-D) agarose gel electrophoresis to map replicons associated with early-replicated, developmental genes. Replicating DNA sequences were radioactively labeled in isolated nuclei obtained from synchronous plasmodia at different times in S phase. DNA purified from nuclei after labeling was then used to probe Southern blots of the battery of cDNAs from various life cycle stages. Hybridization signals indicated at what time in the S phase specific genes were replicated. It was found that 4 of 6 plasmodial genes were replicated by the first 20 min of a 3 hr S phase in the plasmodium, while genes specific for other stages (amoeba, spherule) exhibit later replication than plasmodial genes, at times extending to 60 min into S phase. Two-dimensional agarose gel electrophoresis of restriction digested genomic DNA, isolated at defined times in S phase, indicated close association between transcription units and origins of DNA replication for the three early-replicated, plasmodial genes assayed by this technique. One of these genes, termed LAV1-2, was examined extensively over a 15.2 kb chromosomal region. LAV1=2 was determined to be flanked by origins of replication and to exhibit long-lived, X-shaped structures indicative of replication termination. In addition, a novel replication structure, detected as a previously undescribed hybridization pattern on blots of 2-D gels, was found downstream of LAV1-2 only at very early times in S phase and may be specifically associated with origin activity. Finally, the sequential pattern of replication of LAV1-2 was shown to be identical between alleles of this gene located on different chromosomes. |
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| Item Description: | Typescript (photocopy). Vita. "Major subject: Biology." |
| Physical Description: | xiv, 239 leaves : illustrations ; 29 cm |
| Bibliography: | Includes bibliographical references. |