The replication termination signal terB of the Escherichia coli chromosome is a deletion hot spot.

Hybrids composed of phage M13, plasmid pBR322 and the termination signal of Escherichia coli chromosome replication terB were used to show that arrest of DNA synthesis creates a very efficient deletion hot spot. Up to 80% of deletions occurring in these hybrids had one deletion end-point at terB provided that (i) terB was oriented to arrest M13 and pBR322 leading strand synthesis; and (ii) the host cells contained the Tus protein necessary for arresting DNA synthesis at terB. The position of terB and the flanking sequences had little effect on deletion hot spot activity. About 90% of the deletions at terB ended 5-6 nucleotides in front of the major replication arrest site. We propose two models to account for deletion formation and speculate that many genome rearrangements may be due to the pausing of DNA replication.     

DNA methylation is not increased in mouse-human somatic cell hybrids

The level of DNA methylation in three mouse-human cell lines that retained different human chromosomes and in the parental mouse and human lines has been determined by high-pressure liquid chromatography (HPLC). The level of methylation is similar in the hybrid and parental cells, indicating that interspecific somatic cell hybridization followed by preferential chromosome segregation can occur without an increase in overall DNA methylation.     

Myelogenesis and estimation of the number of axons in the anterior commissure of the chick (Gallus gallus)

Summary By use of light- and electron microscopy the anterior commissure of the chick was studied at different times during development. Between the 19th day of incubation and the 35th day after hatching the cross-sectional area of the anterior commissure, as determined from mid-sagittal sections, undergoes a 6-fold increase in size. Thereafter the area remains fairly constant. The total number of fibres in the anterior commissure was estimated to be 89000. The full complement of fibres is already present by the 19th day of incubation. Myelogenesis occurs mainly between the 19th day of incubation and the 35th day after hatching, concomitant with the increase in cross-sectional area. From the 35th day after hatching, myelinated fibres comprise approximately 40% of the total number of fibres. The median diameter of unmyelinated fibres is about 0.35–0.40 m. The median diameters of myelinated axons and fibres are 0.8–1.0 m and 1.1–1.3 m, respectively.     

Human DNA sequences exhibiting gamete-specific hypomethylation.

Three human DNA sequences have been cloned from DNA regions which are strikingly undermethylated in sperm, highly methylated in adult somatic tissues, and methylated to an intermediate extent in tissues of extraembryonic origin. It is proposed that some such DNA sequences may function specifically early in embryogenesis or during gametogenesis. They may be subsequently extensively methylated in the embryonic cell lineage and methylated to a lesser extent in extraembryonic tissues in order to allow embryogenesis to proceed.     

Molecular cloning and analysis of Staphylococcus aureus chromosomal aminoglycoside resistance genes

Most of the aminoglycoside resistant Staphylococcus aureus strains isolated in France are resistant to all the antibiotics belonging to this family. Two aminoglycoside-modifying enzymes were detected in the wild-type strains studied: an APH3'III and an AAC6'-APH2". These strains also carry two types of streptomycin resistance: high-level resistance due to chromosomal mutation(s) affecting ribosome affinity and low-level resistance, the mechanism of which was not characterized. All the aminoglycoside resistance genes were located on the chromosome. DNA fragments of 1.5 and 1.95 kb carrying the aphA and aacA genes, respectively, were isolated, by cloning, from the cellular DNA of a clinical isolate. When these genes were introduced into Escherichia coli and Bacillus subtilis strains, the enzymes synthesized were indistinguishable from those produced by the S. aureus strains. When the cellular DNAs of wild-type and resistant strains were hybridized with the cloned fragments, sequences homologous to the fragment carrying the aphA gene were found to be located at the same chromosomal site, while those hybridizing with the fragment carrying the aacA gene were at different chromosomal sites.     

Further characterization of cooperative interactions of monoclonal antibodies

We reported previously that mixtures of some monoclonal antibodies directed against the glycoprotein hormone human chorionic gonadotropin (hCG) had a higher affinity for the antigen than either monoclonal antibody separately. The synergistic interaction could no longer be detected when one of the antibodies was replaced with its F(ab) fragment. This cooperative interaction has now been further characterized. One-half of 10 possible pairs prepared from five IgG1 monoclonal antibodies against hCG result in a synergistic interaction. The addition of an IgG2b monoclonal antibody to one of the IgG1 monoclonal antibodies also induces a cooperative interaction, which shows that the effect is not subclass restricted. Cooperative interactions between antibodies are also not restricted to solution conditions; adsorption of one antibody to a solid support appears to increase the cooperative effect. Indeed, one pair of antibodies that failed to bind hCG synergistically in solution did so when one antibody was bound to a solid surface. The liquid phase antibody also has an effect on the specificity of the solid phase antibody. The sensitivity of the solid phase assay system has enabled us to develop a rapid method of determining if two monoclonal antibodies can bind to an antigen simultaneously. A quantitative theoretical model has been devised that successfully predicts the cooperative behavior observed between antibodies and should be useful in devising conditions that result in sensitive solid phase radioimmunoassays.     

Ligation of highly modified bacteriophage DNA

After digestion by TaqI or nicking by DNAase I, five highly modified bacteriophage DNAs were tested as substrates for T4 DNA ligase. The DNAs used were from phages T4, XP12, PBS1, SP82, and SP15, which contain as a major base either glucosylated 5-hydroxymethylcytosine, 5-methylcytosine, uracil, 5-hydroxymethyluracil, or phosphoglucuronated, glucosylated 5-(4′,5′-dihydroxypentyl)uracil, respectively. The relative ability of cohesive-ended TaqI fragments of these DNAs and of normal, λ DNA to be ligated was as follows: $\text{λ}\text{DNA}\text{=}\text{XP}\text{12}\text{DNA}\text{>}\text{SP}\text{82}\text{DNA}\text{?}\text{nonglucosylated}\text{T}\text{4}\text{DNA}\text{>}\text{T}\text{4}\text{DNA}\text{=}\text{PBS}\text{1}\text{DNA}\text{?}\text{SP}\text{15}\text{DNA}$. TaqI-T4 DNA fragments were also inefficiently ligated by Escherichia coli DNA ligase. However, annealing-independent ligation of DNAase I-nicked T4, PBS1, and λ DNAs was equally efficient. We conclude that the poor ligation of TaqI fragments of T4 and PBS1 DNAs was due to the hydroxymethylation (and glucosylation) of cytosine residues at T4's cohesive ends and the substitution of uracil residues for thymine residues adjacent to PBS1's cohesive ends destabilizing the annealing of the restriction fragments. Only SP15 DNA with its negatively charged, modified base was unable to serve as a substrate for T4 DNA ligase in an annealing-independent reaction; therefore, its modification directly interfered with enzyme binding or catalysis.     

Bacteriology of manganese nodules: III. Reduction of MnO(2) by two strains of nodule bacteria

MnO₂ reduction by aerobic growing cultures of Bacillus 29 and coccus 32, isolated from ferromanganese nodules, was assessed for 7 days. A 1-day lag was observed before the onset of MnO₂ reduction by either culture. Addition of HgCl₂ to a final concentration of about 10^-3 M caused a rapid cessation of MnO₂ reduction by the growing cultures. Neither culture reduced MnO₂ when grown under continued anaerobiosis from the start of an experiment. However, if conditions were made anaerobic after MnO₂ reduction was initiated, reduction continued at a rate only slightly lower than that under aerobic conditions. Resting-cell cultures reduced MnO₂ equally well aerobically and anaerobically, provided that ferricyanide was present to serve as electron carrier. These findings showed that oxygen is needed for culture adaptation to MnO₂ reduction, and that oxygen does not interfere with microbial MnO₂ reduction itself. Both cultures caused sharp drops in the pH of the medium during MnO₂ reduction: with coccus 32, during the entire incubation time; with Bacillus 29, for the first 3 days. The E_h of the medium fluctuated with either culture and never fell below 469 mv with Bacillus 29 and below 394 mv with coccus 32. The rates of glucose consumption and Mn²⁺ release by Bacillus 29 and coccus 32 were fairly constant, but the rates of lactate and pyruvate production were not. Although acid production undoubtedly helped in the reduction of pyrolusite (MnO₂) by the bacteria, it did not appear to be important in the reduction of manganese oxide in ferromanganese nodules, as shown by the results with a nodule enrichment.