Please use this identifier to cite or link to this item:
https://doi.org/10.1016/j.ab.2004.03.065
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dc.title | Uracil in DNA, determined by an improved assay, is increased when deoxynucleosides are added to folate-deficient cultured human lymphocytes | |
dc.contributor.author | Mashiyama, S.T. | |
dc.contributor.author | Courtemanche, C. | |
dc.contributor.author | Elson-Schwab, I. | |
dc.contributor.author | Ames, B.N. | |
dc.contributor.author | Crott, J. | |
dc.contributor.author | Fenech, M. | |
dc.contributor.author | Lee, B.L. | |
dc.contributor.author | Ong, C.N. | |
dc.date.accessioned | 2012-04-02T06:54:40Z | |
dc.date.available | 2012-04-02T06:54:40Z | |
dc.date.issued | 2004 | |
dc.identifier.citation | Mashiyama, S.T., Courtemanche, C., Elson-Schwab, I., Ames, B.N., Crott, J., Fenech, M., Lee, B.L., Ong, C.N. (2004). Uracil in DNA, determined by an improved assay, is increased when deoxynucleosides are added to folate-deficient cultured human lymphocytes. Analytical Biochemistry 330 (1) : 58-69. ScholarBank@NUS Repository. https://doi.org/10.1016/j.ab.2004.03.065 | |
dc.identifier.issn | 00032697 | |
dc.identifier.uri | http://scholarbank.nus.edu.sg/handle/10635/31863 | |
dc.description.abstract | Folate deficiency leads to increased dUMP/dTMP ratios and uracil misincorporation into DNA, which may increase cancer risk. We improved a previously described gas chromatography-mass spectrometry (GC-MS) assay for uracil in DNA and validated the assay by analyzing the DNA-uracil content of normal, primary human lymphocytes that were cultured in 0-3000nM folic acid. In addition, the effects of nucleoside mixtures T or TdCA (T, thymidine; A, adenosine; dC, deoxycytidine) were investigated. Over 4 consecutive days, the inter- and intraassay coefficients of variation (CVs) were 2.3-3.9 and 0.6-2.2%. Mean recovery was 99.4%. Oligonucleotides containing 100pg of uracil yielded a mean uracil measurement of 110.1pg (CV=2.7%). Cells grown in different concentrations of folate showed a bimodal response, with maximum DNA-uracil at 12nM, and minima at 0 and 3000nM folate. Extremely folate-deficient cells may incorporate less uracil because DNA synthesis is reduced. A wide response to folate deficiency was seen in cells from different donors, suggesting that genetic background plays a critical role in individual susceptibility to DNA damage and cancer risk. Unexpectedly, TdCA supplementation caused increased DNA-uracil (vs 3000nM folate for 10 days, P<0.05), probably due to the conversion of deoxycytidine to deoxyuridine by cytidine deaminase, leading to elevated dUMP/dTMP ratios. This improved uracil assay could serve as a useful tool in the study of the mechanism of uracil misincorporation into DNA. The assay requires 3μg of DNA per folate-deficient sample, but more may be required for baseline DNA-uracil detection in healthy humans. © 2004 Elsevier Inc. All rights reserved. | |
dc.description.uri | http://libproxy1.nus.edu.sg/login?url=http://dx.doi.org/10.1016/j.ab.2004.03.065 | |
dc.source | Scopus | |
dc.subject | Cultured human lymphocytes | |
dc.subject | DNA damage | |
dc.subject | DNA-uracil detection | |
dc.subject | Folate deficiency | |
dc.subject | Gas chromatography-mass spectrometry | |
dc.type | Article | |
dc.contributor.department | COMMUNITY,OCCUPATIONAL & FAMILY MEDICINE | |
dc.description.doi | 10.1016/j.ab.2004.03.065 | |
dc.description.sourcetitle | Analytical Biochemistry | |
dc.description.volume | 330 | |
dc.description.issue | 1 | |
dc.description.page | 58-69 | |
dc.description.coden | ANBCA | |
dc.identifier.isiut | 000222164700008 | |
Appears in Collections: | Staff Publications |
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