Quantitative reversed-phase high performance liquid chromatographic determination of major and modified deoxyribonucleosides in DNA
TLDR
A method to accurately determine the complete major and modified base composition of a few micrograms of unlabeled DNA in which the m5dCyd comprises only 1 to 2% of the total bases is developed.Abstract:
We have developed a method to accurately determine (< 3% RSD) the complete major and modified base composition of a few micrograms of unlabeled DNA. The DNA samples were quantitatively hydrolyzed with DNase 1, Nuclease P1, and bacterial alkaline phosphatase. The resulting deoxyribonucleosides were directly separated in 70 min by reversed-phase high performance liquid chromatography with detection by ultraviolet absorption at 254 nm and 280 nm (RP-HPLC). The highly sensitive and selective dual wavelength quantitation greatly enhances the precision and accuracy of the chromatographic analysis. Contamination of DNA preparations with RNA does not interfere with the DNA analysis due to the high resolution of the chromatography. We have used this method for the quantitation of m5dCyd in 5 microgram of calf thymus and salmon sperm DNA in which the m5dCyd comprises only 1 to 2% of the total bases. This method should be a useful research tool in studies on various DNAs and DNA subfractions and should help to elucidate the functions of methylation of DNA.read more
Citations
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Precise Measurement of the G+C Content of Deoxyribonucleic Acid by High-Performance Liquid Chromatography
TL;DR: High-performance liquid chromatography is a promising alternative for determining the G+C content of bacterial deoxyribonucleic acid (DNA) and may also be more accurate than indirect methods, such as the buoyant density and thermal denaturation methods.
Journal ArticleDOI
The power and the promise of DNA methylation markers
TL;DR: The past few years have seen an explosion of interest in the epigenetics of cancer as a consequence of both the exciting coalescence of the chromatin and DNA methylation fields, and the realization thatDNA methylation changes are involved in human malignancies.
Journal ArticleDOI
DNMT1 and DNMT3b cooperate to silence genes in human cancer cells.
Ina Rhee,Kurtis E. Bachman,Ben Ho Park,Kam Wing Jair,Ray Whay Chiu Yen,Kornel E. Schuebel,Hengmi Cui,Andrew P. Feinberg,Christoph Lengauer,Kenneth W. Kinzler,Stephen B. Baylin,Bert Vogelstein +11 more
TL;DR: It is demonstrated that two enzymes cooperatively maintain DNA methylation and gene silencing in human cancer cells, and compelling evidence that such methylation is essential for optimal neoplastic proliferation is provided.
Journal ArticleDOI
Amount and distribution of 5-methylcytosine in human DNA from different types of tissues or cells
Melanie Ehrlich,Miguel A. Gama-Sosa,Lan Hsiang Huang,Rose Marie Midgett,Kenneth C. Kuo,Roy A. McCune,Charles W. Gehrke +6 more
TL;DR: Analysis of the total base composition of DNA from seven different normal human tissues and eight different types of homogeneous human cell populations revealed considerable tissue-specific and cell-specific differences in the extent of methylation of cytosine residues.
Journal ArticleDOI
The 5-methylcytosine content of DNA from human tumors
Miguel A. Gama-Sosa,Valerie A. Slagel,Ronald W. Trewyn,Ronald Oxenhandler,Kenneth C. Kuo,Charles W. Gehrke,Melanie Ehrlich +6 more
TL;DR: Most of the metastatic neoplasms had significantly lower genomic m5C contents than did most of the benign neoplasm or normal tissues, which might reflect an involvement of extensive demethylation of DNA in tumor progression.
References
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A simple method for DNA restriction site mapping
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TL;DR: The DNA from two transplantable hepatocellular carcinoma lines was less methylated than predicted rates of cell division in these tumors, suggesting that an aberration in endogenous DNA methylation may occur during neoplastic transformation.
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TL;DR: A positive correlation between the G + C and 5-methylcytosine contents in the DNA of animals belonging to one class is observed and it is believed that the tissue differences with regard to the 5- methylcytose content in theDNA of the organism are due to the various levels and specific character of DNA methylation.
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TL;DR: Since macronuclear DNA from five different syngens of paramecia differed only slightly in their 6 N- methyladenine content, it has been concluded that intersyngen differences can not be attributed to the level of methyl group content.