A preparative method for isolating high-molecular-weight DNA from animal cells is described. This method is based on the use of proteinase K, a powerful proteolytic enzyme with a broad action spectrum, which is very active in the presence of sodium dodecylsulfate and ethylene-diamine tetraacetate. The DNA preparation is free of RNA, protein and degrading enzymes. The number-average molecular weight of the native DNA is 190 × 106, whereas it is 90 × 106 for single-stranded DNA, indicating that the DNA molecules do not contain single-stranded nicks. The native DNA molecules range in molecular weight from 40 × 106 to more than 500 × 106.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1973.tb02881.x

Isolation of high-molecular-weight DNA from mammalian cells.

On the mechanism of DNA replication in mammalian chromosomes

Postreplication repair of DNA in ultraviolet-irradiated mammalian cells

Since their introduction (Heidelberger et al., 1957), the fluorinated pyrimidines and their nucleosides have been widely used as biochemical tools for the elucidation of a number of problems encountered in cell biology and molecular biology. Of more practical importance, however, has been their extensive use as drugs for the palliative treatment of patients suffering from disseminated cancer. One of the compounds is now clinically useful in the curative treatment of herpes simplex viral infections of the eye. Although these compounds have generated an enormous literature, this chapter will deal primarily with a review of the major pharmacological and biochemical effects of these compounds; the coverage of the literature, by necessity, will focus only on key references that are germane to the topics being described. Several reviews of this topic have appeared elsewhere (Heidelberger and Ansfield, 1963; Heidelberger, 1965, 1966, 1967, 1969, 1970, 1973; Mandel, 1969; Carter, 1970).

Fluorinated Pyrimidines and Their Nucleosides

• A general model for the production of aberrations in eukaryote chromosomes is proposed. The basic assumptions of this model are: • (1) The chromosome is mononeme, containing but a single DNA double helix before replication; (2) Aberrations result only from lesions in DNA; (3) The primary lesions may be either polynucleotide strand breaks or altered bases which are converted to strand breaks either through the operation of enzymatic repair mechanisms or as a consequence of normal semiconservative DNA replication; (4) Final expression of polynucleotide strand breaks as chromosomal aberrations of various types is the consequence of the operation of repair mechanisms, DNA replication, and a single strand DNAase. • It is shown that this model is consitent with the available data on chromosomal aberration production by a variety of chemical agents as well as by ionizing radiation. In particular, it accounts for the observed differences in the types of aberrations produced by these two classes of aberration producing agents at different points in the cell cycle.

Mechanisms of chromosomal aberration production III. Chemicals and ionizing radiation

Radiosensitivity of cells of Chinese hamster in vitro in relation to the cell cycle.

Evaluation of S phase synchronization by analysis of DNA replication in 5-bromodeoxyuridine.

RECENT evidence indicates that the response of Chinese hamster cells in vitro to ultra-violet radiation changes during the difference phases of the cell cycle1,2,4. Humphrey et al.1 have shown that cells synthesizing deoxyribo-nucleic acid (DNA) at the time of irradiation using ultraviolet rays (S phase cells) sustain more chromosome damage than cells which were in the period before DNA synthesis (G1 or cells in the period after DNA synthesis (G2). Sinclair and Morton have demonstrated that the response to ultra-violet radiation in synchronous populations of cells as measured by cell death also fluctuates with the age of the irradiated cells; the fraction of surviving cells falls as the cells move from G1 into S phase, and rises as they move from S into G2 phase2. In an effort to explain these changes in radiation response during the cell cycle, the production of an ultra-violet photoproduct of DNA, the thymine dimer3 has been measured in synchronized population of Chinese hamster cells4.

Ronald M. Humphrey

Papers

Radiosensitivity of cells of Chinese hamster in vitro in relation to the cell cycle.

Evaluation of S phase synchronization by analysis of DNA replication in 5-bromodeoxyuridine.

Induction of thymine dimers in synchronized populations of Chinese hamster cells.

The cell-cycle phase synthesis of non-histone proteins in mammalian cells.

Responses of Chinese Hamster and L Cells to 2′-Deoxy-5-Fluoro-Uridine and Thymidine