A new DNA-cloning vector for Haemophilus influenzae Rd

A new, high-efficiency, DNA-cloning vector pJ1-8 was derived in two steps from the chimeric plasmid pD7 consisting of RSF 0885(ampr) and Haemophilus influenzae chromosomal DNA. pJl-8 has only one EcoRI site and a molecular weight of only 2.5×106. No detectable ampr transformation was obtained with pJl-8 DNA. However, ampr transformation increases markedly if Haemophilus influenzae chromosomal DNA segments are spliced into it, providing a very facile assay for detecting inserts

Analysis of a genetically unstable region in Streptomyces lividans 66-TK64

Genetic and molecular analyses of an unstable region encompassing the gene loci cml arg and a 5.7 kb amplifiable unit of DNA were done. Spontaneous mutants from Cm1R→CmlS and the revertants from CmlS →CmlR were analysed for mutations at arg locus and amplification of amplifiable unit of DNA. Twenty-one revertants were analysed. Two of these had large-scale amplification and one of these was also Arg-. Nine of the revertants which were Arg+ had low-level or intermediate-level amplification of the 5.7 kb DNA sequence but no deletions of the flanking sequences were detected. Five of the CmIR' revertants, which were also Arg+, had lost one of the two copies from the doublet of amplifiable unit of DNA. The remaining five revertants did not show any other change. The amplifiable unit of DNA, therefore, not only undergoes amplification but can also suffer specific deletion of one copy. Thus, this region as a whole is characterized by instability and the events appear to take place at more than one locus concomitantly with a high frequency

Genetic transformation in bacteria

Certain species of bacteria can become competent to take up high molecular weight DNA from the surrounding medium. DNA homologous to resident chromosomal DNA is transported, processed and recombined with the resident DNA. There are some variations in steps leading to transformation between Gram-positive bacteria likebiplococcus pneumoniae and Gram-negative bacteria represented byHaemophilus influenzae but the integration is by single-strand displacement in both cases. Plasmid (RSF0885) transformation is low inHaemophilus influenzae but this is increased significantly if (homologous) chromosomal DNA is spliced to plasmid DNA. In Haemophilus influenzae, rec1 function is required for peak transformation with chimeric plasmids. Chimeric plasmid fixed presumably extrachromosomally undergoes frequent recombination between homologous segments contained in resident chromosome and the plasmid

Transformation of Haemophilus influenzae by plasmid RSF0885

Plasmid RSF0885, which conferred ampicillin resistance, transformed competent Haemophilus influenzae cells with low efficiency (maximum, less than 0.01%). As judged by competition experiments and uptake of radioactivity, plasmid RSF0885 deoxyribonucleic acid was taken up into competent H. influenzae cells several orders of magnitude less efficiently than H. influenzae chromosomal deoxyribonucleic acid. Plasmid RSF0885 transformed cells with even lower efficiency than could be accounted for by the low uptake. Transformation was not affected by rec-1 and rec-2 mutations in the recipient, and strains cured of the plasmid did not show increased transformation. Plasmid molecules cut once with a restriction enzyme that made blunt ends did not transform. Transformation was favored by the closed circular form of the plasmid

An estimate of the physical distance between two linked markers in Haemophilus influenzae

Using DNA clones, the physical distance between the linked genesnov and str in Haemophilus influenzae was estimated. Although none of the cloned inserts contained both the markers, pJ1-8StrR 13 (insert of 18·7 kb) included str gene at one end and part of nov gene at the other end of the insert. By EcoRI restriction analysis and by Southern hybridization, the distance between the two EcoRI sites, cutting at which inactivates the two genes, was estimated to be 17·7 kb. A single continuous EcoRI fragment (containing 4EcoRI sites within it) carrying both the genes intact would need to be 20·4 kb in size. These estimates were confirmed independently using different clones of novr and strr alleles as probes for hybridization with BamHI-digested chromosomal DNA

Coulomb and nuclear breakup of a halo nucleus 11Be

Breakup reactions of the one-neutron halo nucleus 11Be on Pb and C targets at about 70 MeV/u have been investigated by measuring the momentum vectors of the incident 11Be, outgoing 10Be, and neutron in coincidence. The relative energy spectra as well as the angular distributions of the 10Be+n center of mass have been extracted for both targets. For the breakup on Pb target, the selection of forward scattering angles is found to be effective to extract almost purely the first-order E1 Coulomb breakup component, and to exclude the nuclear contribution and higher-order Coulomb breakup components. This angle-selected energy spectrum is thus used to deduce the spectroscopic factor for the 10Be(0+) 2s_1/2 configuration in 11Be which is found to be 0.72+-0.04 with B(E1) up to Ex=4 MeV of 1.05+-0.06 e2fm2. The energy weighted E1 strength up to Ex=4 MeV explains 70+-10% of the cluster sum rule, consistent with the obtained spectroscopic factor. The non-energy weighted sum rule is used to extract the root mean square distance of the halo neutron to be 5.77(16) fm, consistent with previously known values. In the breakup with C target, we have observed the excitations to the known unbound states in 11Be at Ex=1.78 MeV and 3.41 MeV. Angular distributions for these states show the diffraction pattern characteristic of L=2 transitions, resulting in J^pi =(3/2,5/2)+ assignment for these states. We finally find that even for the C target the E1 Coulomb direct breakup mechanism becomes dominant at very forward angles.Comment: 14 pages, 7 figures, accepted for publication on Physical Review

Monohybrid and dihybrid segregations in the progenies of tobacco transformed for kanamycin resistance with a Ti-vector system

A chimeric DNA construction having nopaline synthase promoter, coding sequences of neomycin phosphotransferase gene conferring resistance to antibiotic kanamycin and OCS (octopine synthase) polyadenylation sequences bracketed by T-DNA ends was transferred to tobacco. Leaf discs were infected with A. tumefaciens containing disarmed, cointegrate plasmid pGV3850:: 1103 and allowed to form a callus in the presence of kanamycin. Shoots regenerated from infected leaf discs either through the callus or arising directly were further selected for their ability to root in kanamycin-containing media. Among the nine transgenic plants that were progeny tested, the transferred bacterial gene segregated as monohybrid ratio (3 KanR: 1 KanS) in seven. Segregation data of two plant progenies indicated the presence of two independent loci of KanR DNA insertion (15 KanR: 1 KanS). Back-cross segregation data were consistent with the monohybrid or independent assortment of duplicate factors. Thus in the two cases, a minimum independent integration of two copies of T-DNA each with a KanR marker is inferred

Projectile fragmentation reactions and production of nuclei near the neutron drip-line

The reaction mechanism of projectile fragmentation at intermediate energies has been investigated observing the target dependence of the production cross sections of very neutron-rich nuclei. Measurement of longitudinal momentum distributions of projectile-like fragments within a wide range of fragment mass and its charge was performed using a hundred-MeV/n $^{40}$Ar beam incident on Be and Ta targets. By measurement of fragment momentum distribution, a parabolic mass dependence of momentum peak shift was observed in the results of both targets, and a phenomenon of light-fragment acceleration was found only in the Be-target data. The analysis of production cross sections revealed an obvious enhancement of the target dependence except target size effect when the neutron excess is increased. This result implies the breakdown of factorization (BOF) of production cross sections for very neutron-rich nuclei near the drip line.Comment: 16 pages, 18 figures, submitted to Phys. Rev.