The esperamicins represent a class of antitumor antibiotics characterized by an unusual chemical core structure and extremely potent cytotoxicity. The mechanism by which these drugs produce cytotoxicity was investigated and found to be related to the formation of single- and double-strand DNA breaks. Using five structurally related analogs, we defined a structure-activity relationship for cytot...

Genes encoding galactose utilization in Vibrio cholerae have been cloned into Escherichia coli host strains in order to determine their functions and organization. BarnHI-digested chromosomal DNA from 01 V. cholerae strain 26-3 was ligated into the cloning vector pACYC 184. A chimeric plasmid, pHS17, containing a 5.9-kb insert expressed galactokinase activity in a gaK mutant of E. coli. However...

The ebgA (evolved beta-galactosidase) gene of Escherichia coli was isolated as part of a 9.6-kilobase (kb) sequence cloned into plasmid pBR322. The position of the ebgA gene within that 9.6-kilobase sequence was identified by insertional inactivation by means of the transposon gamma-delta. In addition to the gamma-delta insertions that inactivate ebgA by disrupting the coding sequence, seven ad...

The Escherichia coli uvrB gene, located on a 1.5-megadalton EcoRI (fragment F, derived from transducing phage lambda b2att2 [lambda b2cI857intam6 delta (bioAB)bio-FCD+uvrB+], has been cloned in the unique EcoRI site of several "relaxed" plasmids, i.e., pMB9, pBR322, and pBH20 (= ;BR322, including the lac regulatory elements [K. Itakura, T. Hirose, R. Crea, A. D. Riggs, H. L. Heyneker, F. Boliva...

Genomic DNA from Streptococcus equisimilis strain H46A was cloned in Escherichia coli by using the bacteriophage lambda replacement vector L47 and an in vitro packaging system. A casein/plasminogen overlay technique was used to screen the phage bank for recombinants carrying the streptokinase gene ( skc ). The gene was present with a frequency of 1 in 836 recombinants, and 10 independent clones...

In the presence of O2 and 5 mM dithiothreitol, penta-N-methylpyrrolecarboxamide-EDTA-Fe(II) [P5E*Fe(lI)] at 0.5 ILM cleaves pBR322 plasmid DNA (50 pAM in base pairs) on opposite strands to afford discrete DNA fragments as analyzed by agarose gel electrophoresis. High-resolution denaturing gel electrophoresis of a nP-end-labeled 517-base-pair restriction fragment containing a major cleavage site...

Ionizing radiation induces a variety of DNA damages including single-strand breaks (SSBs), double-strand breaks (DSBs), abasic sites, modified sugars, and bases. Most theoretical and experimental studies have been focused on DNA strand scissions, in particular production of DNA double-strand breaks. DSBs have been proven to be a key damage at a molecular level responsible for the formation of c...

The definition of the functional boundaries of regulatory DNA elements often includes sequential deletion of DNA using controlled digestion with BaBl nuclease [for example, (1, 2)]. To obtain small deletions (< 100 bp) we have devised a simple strategy involving cloning of the target DNA into the EcoSl site of the pBR322 plasmid. The hybrid plasmid is linearized at a unique restriction site pre...

A recombinant DNA plasmid, pBR322-GH1, which contains about 80% of the sequences of rat pregrowth hormone (pGH) mRNA, allowed an analysis of nuclear RNA from GH3 cells for possible precursors of cytoplasmic pGH mRNA. A single 20-22S RNA SPECIES ABOUT 2-3 TIMes larger than pGH mRNA was detected in nuclear RNA from GH3 cells labeled for 5 min. with 3H-uridine. After longer label times a 12S RNA i...

Two cationic γ-carbolines, 2-methyl-5H-pyrido[4,3-b]indolium iodide (MPII) and 2,5-dimethyl-5H-pyrido[4,3-b]indolium iodide (DPII), were synthesized, and the DNA-binding properties of the cationic γ-carbolines were elucidated. Through a series of experiments, we proved that the two cationic γ-carbolines could strongly interact with DNA by intercalative binding. However, DPII, with a methyl grou...