Detection of single-base mutations in a mixed population of cells: A comparison of SSCP and direct sequencing |
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| Affiliation: | 1. Laboratory of Environmental Pollution Control, Department of Chemistry, Aristotle University of Thessaloniki, GR–541 24 Thessaloniki, Greece;2. Centre for Interdisciplinary Research and Innovation (CIRI-AUTH), Balkan Center, Thessaloniki, 10th km Thessaloniki-Thermi Rd, GR 57001, Greece;3. Laboratory of Polymer and Colors Chemistry and Technology, Department of Chemistry, Aristotle University of Thessaloniki, Thessaloniki, GR-541 24, Greece;4. EYATH S.A., Thessaloniki Water Supply & Sewerage Company, Egnatias 127, GR 54635, Thessaloniki, Greece;5. Department of Chemistry, International Hellenic University, Kavala GR-654 04, Greece;1. Department of Zoology, University of Lucknow, Lucknow, U.P. 226007, India;2. Department of Zoology, Babasaheb Bhimrao Ambedkar University, Lucknow, U.P. 226025, India;1. Department of Microbiology and Immunology at The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Australia;2. Microbiological Diagnostic Unit Public Health Laboratory, Department of Microbiology and Immunology at The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Australia;3. Communicable Disease Epidemiology and Surveillance, Health Protection Branch, Public Health Division, Department of Health, Melbourne, Australia;4. Mycobacterium Reference Laboratory, Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital at The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Australia;5. Royal Melbourne Hospital, Melbourne, Australia;6. Victorian Tuberculosis Program. Melbourne Health at the Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Australia;7. Department of Infectious Diseases at The Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Australia;8. Centre for Pathogen Genomics, University of Melbourne, Australia |
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| Abstract: | Single-base-pair changes in several genes have been shown to be important in carcinogenesis. We have compared the sensitivity of two commonly used techniques for detection of single-base-pair changes. Defined populations of cells were prepared by mixing wild-type Chinese hamster ovary cells with cells carrying a known mutation in the CTP synthetase gene. RNA was extracted and analyzed for the mutation by single-strand conformational polymorphism and direct sequence analysis of polymerase chain reaction products. We found that both techniques were able to detect the mutation when it was present in 25% of the cells, but that direct sequencing was slightly more sensitive and able to detect the mutation when it was present in only 20% of the population. |
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