IMMORTALIZATION AND CHARACTERIZATION OF HUMAN FIBROBLASTS TRANSFORMED WITH AN ORI(-) SV40 GENOME.
Item
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Title
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IMMORTALIZATION AND CHARACTERIZATION OF HUMAN FIBROBLASTS TRANSFORMED WITH AN ORI(-) SV40 GENOME.
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Identifier
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AAI8614695
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identifier
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8614695
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Creator
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NEUFELD, DAVID SAMUEL.
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Contributor
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Harvey L. Ozer
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Date
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1986
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biology, Molecular
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Abstract
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Human diploid cells derived from normal tissue undergo a characteristic pattern of growth in cell culture, ending in a non-replicative phase (Phase III), and, finally, cell death. Human diploid fibroblasts infected in vitro by the oncogenic DNA virus, SV40, become immortal at an enhanced frequency. Considerable viral DNA synthesis is also evident. To preclude viral DNA replication, HS74BM (a normal human diploid fibroblast) was transformed with an ori('-) mutant of SV40. Such ori('-) transformed human fibroblasts have an extended lifespan in culture, beyond that seen in human fibroblasts transformed with a viral DNA (ori('+)), but eventually also senesce. However, a number of continuous cell lines were isolated with the ori('-) transformed human fibroblast. This could not be duplicated with ori('+) transformed human fibroblast. Three cell lines were obtained by blind passaging the cells and picking colonies which grew out of mass cultures. Another three cell lines were isolated by performing a cloning experiment. I then investigated the SV40 integrants, and found only one or two integrants for each line, one of which had apparently been conserved in all the immortalized lines. I concentrated my efforts on Cl-39, which contained only one integrant. Southern analysis was performed to characterize the integrated sequences. Most of the late region and part of the early region were found to be missing. Consistent with absence of part of the early region, a truncated large T antigen was found.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
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Program
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Biology
