Molecular Dosimetry Studies of Forward Mutation Induced at the YG2 Locus in Maize by Ethyl Methanesulfonate
Schy, William Emanuel
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https://hdl.handle.net/2142/77616
Description
Title
Molecular Dosimetry Studies of Forward Mutation Induced at the YG2 Locus in Maize by Ethyl Methanesulfonate
Author(s)
Schy, William Emanuel
Issue Date
1987
Doctoral Committee Chair(s)
Plewa, Michael J.
Department of Study
Genetics and Development
Discipline
Genetics and Development
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Genetics
Language
eng
Abstract
The yg2 assay in Zea mays detects forward mutation in leaf primordia of developed embryos. Studies on the molecular dosimetry of ethyl methanesulfonate (EMS) were conducted using this assay. Parallel genetic and molecular dosimetry experiments were conducted in which the frequency of forward mutation and the level of covalently bound ethyl DNA adducts were determined. Prepared kernels were treated for 8 h at 20$\sp\circ$C with 1 to 10 mM EMS. EMS yielded a concentration-dependent increase in mutation induction proportional to the exposure concentration. The kinetics of mutation induction demonstrated in the intact maize system were consistent with the kinetics observed in earlier in vitro model systems using cultured mammalian cells, and contrasted with the exponential increase in mutation induction characteristic of microbial species. Parallel molecular dosimetry experiments were conducted under identical treatment conditions except using ($\sp3$H) EMS. DNA was extracted and purified from embryonic tissues containing the leaf primordia, the target tissue of the yg2 assay. DNA was quantified and its purity was confirmed by combining two independent methods of DNA determination, UV spectrophotometry and fluorescence measurement following reaction of hydrolyzed DNA with 3,5-diaminobenzoic acid dihydrochloride (DABA). The increase in molecular dose measured was slightly less than proportional to the exposure concentration of EMS. Using concentration as a common parameter between the parallel genetic and dosimetry studies, mutation induction appeared to increase nearly in direct proportion to the molecular dose. However, studies in other genetic systems indicate that the levels of specific DNA adducts, such as O$\sp6$-ethylguanine (O$\sp6$-EtGua) show a better correlation with mutation induction than molecular dose. Neither molecular dose, nor O$\sp6$-EtGua levels account for differences in the absolute frequencies of mutation induction observed in different genetic systems. Therefore, reliable assessment of health risks posed to humans by chemical mutagens appears to require consideration of other factors in addition to DNA dose or adduct formation, including differences in repair capabilities and in the size of the genetic targets in humans relative to the model genetic systems under study.
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