Poplar (Populus nigra L.) plants transformed with aBacillus thuringiensis toxin gene: insecticidal activity and genomic analysis

Insect-resistant poplar (Populus nigra L.) plants have been produced by infecting leaves withAgrobacterium tumefaciens strains carrying a binary vector containing different truncated forms of aBacillus thuringiensis (B.t.) toxin gene under a duplicated CaMV 35S promoter. Putative transgenic plants were propagated by cuttings at two experimental farms (in Beijing and Xinjiang, China). At 2–3 years after transformation, 17 of them were selected on the bases of insect-tolerance and good silvicultural traits, and evaluated for insect resistance, for the presence of theB.t. toxin DNA fragment (Southern blots and PCR) and for the expression of the transgene (western and northern blots). Somaclonal variation, as suggested by the appearance of permanent changes in the shape of the leaves, was also investigated with molecular tools (RFLP (restriction fragment length polymorphism), RAPD (random amplified polymorphic DNA) and microsatellite DNA).Bioassays withApochemia cineraius andLymantria dispar on the leaves of the selected clones showed different and, in some cases, high levels of insecticidal activity. The molecular analysis demonstrated integration and expression of the foreign gene. Somatic changes were correlated to extensive genomic changes and were quantified in dendrograms, in terms of genomic similarity. The analysis of control plants suggested that genomic changes were correlated to thein vitro culture step necessary forA. tumefaciens-mediated gene transfer, rather than to the integration of the foreign genes.Three transgenic clones (12, 153 and 192), selected for insect resistance, reduced morphological changes and promising silvicultural traits, are now under large-scale field evaluation in six different provinces in China.     

Expression, Purification, and Initial Characterization of the Recombinant Storage Protein Precursor ofTheobroma cacao

The gene encoding the 67-kDa cocoa storage protein precursor has been cloned fromTheobroma cacaoand expressed inEscherichia coliusing the pET expression system. The recombinant storage protein has been renatured from inclusion bodies at 30°C using 20 m glycine–NaOH buffer, pH 10.0, containing 1 m oxidized glutathione and 0.1% Brij. The renatured protein was purified and demonstrated to adopt a stable native conformation by optical spectroscopy. Secondary structure analysis from circular dichroism indicated the protein to be 23% α-helix and 38% β-sheet, in close agreement with values obtained using a secondary structure prediction program.     

An ultra-pure in vitro phase synchrony method employing centrifugal elutriation and viable flow cytometric cell sorting

We present a method of synchronizing cells in G1-, S-, and G2M-phases employing sequential centrifugal elutriation and viable flow cytometric cell sorting of Hoechst-33342 stained Chinese hamster ovary cells. G1- and S-phase cells can be separated to greater than 99% homogeneity and G2-M to 70% purity. Most of the 30% contamination in the G2-M fraction was due to S-phase cells, whose reproductive integrity could be eliminated through the use of high specific activity 3H-TdR. There were minimal toxic effects or perturbations to growth following the selection procedures. The most significant limitation of this technique appears to be the rate of cell sorting, which, with current equipment, is approximately 3,000 cells per second.     

Shake Flask Biodegradation of 14 Commercial Phthalate Esters

An acclimated shake flask CO₂ evolution test was used to study the biodegradability of 14 commercial phthalate esters that are commonly used as plasticizers. Both CO₂ evolution (ultimate biodegradation) and loss of parent phthalate esters (primary biodegradation) were measured. With only a few exceptions, primary biodegradation was 90% or higher, and ultimate biodegradation was in excess of 55% of theoretical results in 28 days. The results showed that all of the commercial phthalate esters were susceptible to biodegradation by mixed populations of microorganisms from natural sources. The results also provide considerable insight into the utility and reproducibility of a standard biodegradation test that is being recommended for widespread screening of chemicals.     

The contributions of diverse sense organs to the control of leg movement by a walking insect

Summary During locomotion, stick insectsCarausius morosus, place the tarsus of the rear leg near the tarsus of the ipsilateral middle leg, whatever the position of the latter. This adjustment by the hind leg requires that it receive information on the actual position of the middle leg tarsus. It is shown by ablation experiments that such information is contributed by the following proprioceptors of the middle leg: the ventral and dorsal coxal hairplates, the coxal hair rows, the trochanteral hairplate and the femoral chordotonal organ. Additional information comes from other, as yet unidentified, sense organs. Several alternatives are considered to explain how the signals from the diverse sense organs of the subcoxal joint might be combined in computing the target position for the protracting hind leg. The experimental results support the hypothesis that the signals are added nonlinearly and that a signal deviating from the majority pattern is weighted less.Abbreviations cxHPu ventral coxal hairplate - cxHPd dorsal coxal hairplate - trHP trochanteral hairplate - HR hair row - feCO femoral chordotonal organ - AEP anterior extreme position     

Characterization of temperate phages ofBacillus subtilis

Nine known temperature phages ofBacillus subtilis, including four that are newly isolated (ϱ6, ϱ10, ϱ14, and ϱ18), have been compared. Analysis by serology, immunity, host range, and adsorption site similarity place the phages into four groups: Group I, ϕ105, ϱ6, ϱ10, and ϱ14, which are 80–90% related; Group II, SPO2; Group III, ϕ3T and ϱ11, 100% related; and Group IV, SP16. The phage ϱ18 is largely uncharacterized, but is heteroimmune to other groups.     

Decreased binding of cytotoxic antibody by developing Schistosoma mansoni. Evidence for a surface change independent of host antigen adsorption and membrane turnover.

Schistosoma mansoni schistosomula maintained in chemically defined culture media became increasingly resistant to the cytotoxic effects of infected guinea pig serum. Two- and 6-day-old schistosomula recovered from mice showed no uptake of IgG antibody from infected guinea pig serum, as revealed by the indirect fluorescent antibody test. Results from this test remained negative when the schistosomula were tested at 0 C, after exposure to drugs which inhibit synthetic and secretory processes, or after being killed by heat or formalin. In contrast, new schistosomula collected within 3 hr after skin penetration bound IgG from infection serum under all test conditions, and showed increased susceptibility to cytotoxicity after exposure to various drugs. It thus appears that soon after skin penetration schistosomula undergo surface changes which prevent binding of antibody from infection serum. These changes can apparently take place in the absence of host antigens, and once they have occurred, do not depend on worm physiological processes for their function.     

Electrostatic components of drug-receptor recognition. I. Structural and sequence analogues of DNA polynucleotides

The electrostatic fields associated with the important biological receptor DNA have been studied by means of stereoscopic displays to investigate drug-receptor recognition processes. This revealed great differences between A- and B-type structures and enabled significant nucleotide sequence effects to be detected for the latter helix. These variations were further investigated by topological analysis of the surface potential in the two grooves of the B-DNA duplex at different radii from the helix axis. This made it possible to characterize the potential surface and to allocate curvature changes to specific atomic groupings. A general finding was that larger potential fields were found in the space encompassed by the narrow groove with strong potential gradients from the ends of the helix to the centre in both grooves. This gradient may provide a motive force for translating small molecules on the surface of a polynucleotide.