Measurement of telomeric DNA content in human tissues.

Telomeres, nucleoprotein complexes at the ends of eukaryotic chromosomes, are 10-12 kbp in length in somatic cells, but as small as 1-2 kbp in rapidly growing cancer cells. Southern blot analysis is currently the standard method for the measurement of telomere length. However, accurate determinations are not possible when DNA is broken or scant. To avoid these problems, a slot blot assay that quantitates the relative content, instead of length, of telomere DNA was developed. The relative contents of telomere DNA determined by this slot blot assay were directly proportional to the relative lengths of telomere DNA determined in parallel by Southern blot analysis. Relative telomere DNA content could be measured in samples containing as little as 15 ng of total DNA. Relative telomere DNA content, but not length, also was unaffected by breakage of DNA into fragments 1 kbp or less in length.

Structure and function of the class C tetracycline/H+ antiporter: three independent groups of phenotypes are conferred by TetA (C).

The class C tetracycline/H+ antiporter, TetA(C), confers nine distinct phenotypes in Escherichia coli: resistance to tetracycline, reduced culture density at stationary phase (growth yield), increased supercoiling of plasmid DNA, delayed growth in succinate minimal medium, complementation of potassium uptake defects, increased susceptibility to cadmium, increased susceptibility to fusaric acid, increased susceptibility to bleomycin and increased susceptibility to several classes of cationic aminoglycoside antibiotics. These nine phenotypes were resolved into three 'linkage' groups based on their patterns of suppression by mutations of the tetA(C) gene of plasmid pBR322. Group I includes resistance to tetracycline, increased susceptibility to cadmium and reduced growth yield. Group II includes delayed growth in succinate minimal medium and complementation of potassium uptake defects. Group III includes increased supercoiling of plasmid DNA and increased susceptibilities to fusaric acid, bleomycin and cationic aminoglycosides. Phenotypes of Groups II and III, but not Group I, also were conferred by a chimeric gene encoding a fusion between the N-terminal 34 residues of TetA(C) and the C-terminal 429 residues of a structurally-similar protein, the E. coli galactose/H+ symporter, GalP. In contrast, none of these phenotypes was conferred by a chimeric gene encoding a fusion between the N-terminal 34 residues of TetA(C) and a structurally-dissimilar protein, TEM beta-lactamase. These results demonstrate that the three groups of linked phenotypes are dependent on different elements of the TetA(C) amino acid sequence, implying that TetA(C) confers these phenotypes by at least three independent mechanisms.