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Title: Molecular analysis of mutation in mammalian cells
Author: Shamsher, M. K.
Awarding Body: University College of Swansea
Current Institution: Swansea University
Date of Award: 1994
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Most of the currently available mammalian mutation systems depend on the selection of a mutant phenotype. These selective systems include resistance to the effects of toxic drugs, as is widely used in various mammalian cultured cell systems, and the outgrowth of mutant cells. These systems have served us well for many years but they have their limitations, especially as they can only be used to detect selected target genes in the genome. Here, the restriction site mutation (RSM) assay is proposed as a methodology for the detection of DNA base changes in restriction enzyme recognition sites, without the need for prior selection methods. Changes within a restriction site, created by base mutation renders an inherent restriction site on target genomic DNA resistant to digestion with the particular endonucleases. These unrestricted sequences are then amplified with the polymerase chain reaction where enrichment of the mutated sequence occurs. The developmental stages of the RSM, its applications and limitations are described here when applied to the study of point mutations in the ras oncogene from various sources, namely tissue culture cells, vectors and various tissues from intact mammals. The relative ease and high sensitivity of this technique makes it a suitable method for identifying mutations at specific sites in DNA, especially when the mutated target sequences are present at low frequencies. Here. the methodology has been shown to be applicable to any species for which the DNA sequence information is available.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available