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Quantitative analysis of nucleic acid alkylation in vitro and in cell culture

Thesis/Dissertation ·
OSTI ID:5776204
Chemical modification of polycytidylic acid with methyl methane-sulfonate has been studied by /sup 13/C NMR of the intact polymer, /sup 1/H NMR of the polymer, and /sup 1/H NMR, high-pressure liquid chromatography, and secondary-ion mass spectrometry (SIMS) of the nucleosides following enzymatic degradation. Using synthesized (/sup 2/H/sub 3/-methyl) labeled methylated nucleosides as internal references, the comparison of the relative amounts of products formed when calf thymus DNA is reacted with the weak carcinogen, methyl methanesulfonate and the potent carcinogen, 1-methyl-1-nitrosourea is achieved. Extension of this methodology to in vivo systems requires the recovery of highly purified DNA from the cells for accurate total methylation determinations by /sup 14/C incorporation which is achieved here by enzymatic and hydroxyapatite purifications. Upon alkylation of V79 Chinese hamster cells with methyl methanesulfonate, again only one methyl product, 7-methyl-2'-deoxyguanosine is detected.
Research Organization:
Purdue Univ., Lafayette, IN (USA)
OSTI ID:
5776204
Country of Publication:
United States
Language:
English

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