ALBERT

Notes: Abstract. We examined the structure, intranuclear distribution and activity of ribosomal DNA (rDNA) in Nico-tiana sylvestris (2n=2x=24) and N. tomentosiformis (2n=2x=24) and compared these with patterns in N. tabacum (tobacco, 2n=4x=48). We also examined a long-established N. tabacum culture, TBY-2. Nicotiana tabacum is an allotetraploid thought to be derived from ancestors of N. sylvestris (S-genome donor) and N. tomentosiformis (T-genome donor). Nicotiana sylvestris has three rDNA loci, one locus each on chromosomes 10, 11, and 12. In root-tip meristematic interphase cells, the site on chromosome 12 remains condensed and inactive, while the sites on chromosomes 10 and 11 show activity at the proximal end of the locus only. Nicotiana tomentosiformis has one major locus on chromosome 3 showing activity and a minor, inactive locus on chromosome 11. In N. tabacum cv. 095-55, there are four rDNA loci on T3, S10, S11/t and S12 (S11/t carries a small T-genome translocation). The locus on S12 remains condensed and inactive in root-tip meristematic cells while the others show activity, including decondensation at interphase and secondary constrictions at metaphase. Nicotiana tabacum DNA digested with methylcytosine-sensitive enzymes revealed a hybridisation pattern for rDNA that resembled that of N. tomentosiformis and not N. sylvestris. The data indicate that active, undermethylated genes are of the N. tomentosiformis type. Since S-genome chromosomes of N. tabacum show rDNA expression, the result indicates rDNA gene conversion of the active rDNA units on these chromosomes. Gene conversion in N. tabacum is consistent with the results of previous work. However, using primers specific for the S-genome rDNA intergenic sequences (IGS) in the polymerase chain reaction (PCR) show that rDNA gene conversion has not gone to completion in N. tabacum. Furthermore, using methylation-insensitive restriction enzymes we demonstrate that about 8% of the rDNA units remain of the N. sylvestris type (from ca. 75% based on the sum of the rDNA copy numbers in the parents). Since the active genes are likely to be of an N. tomentosiformis type, the N. sylvestris type units are presumably contained within inactive loci (i.e. on chromosome S12). Nicotiana sylvestris has approximately three times as much rDNA as the other two species, resulting in much condensed rDNA at interphase. This species also has three classes of IGS, indicating gene conversion has not homogenised repeat length in this species. The results suggest that methylation and/or DNA condensation has reduced or prevented gene conversion from occurring at inactive genes at rDNA loci. Alternatively, active undermethylated units may be vulnerable to gene conversion, perhaps because they are decondensed and located in close proximity within the nucleolus at interphase. In TBY-2, restriction enzymes showed hybridisation patterns that were similar to, but different from, those of N. tabacum. In addition, TBY-2 has elevated rDNA copy number and variable numbers of rDNA loci, all indicating rDNA evolution in culture.

Notes: Abstract. Phylogenetic schemes based on changing DNA sequence have made a major impact on our understanding of evolutionary relationships and significantly built on knowledge gained by morphological and anatomical studies. Here we present another approach to phylogeny, using fluorescent in situ hybridisation. The phylogenetic scheme presented is likely to be robust since it is derived from the chromosomal distribution of ten repetitive sequences with different functions and evolutionary constraints [GRS, HRS60, NTRS, the Arabidopsis-type telomere repeat (TTTAGGG)n, 18S-5.8S-26S ribosomal DNA (rDNA), 5S rDNA, and four classes of geminiviral-related DNA (GRD)]. The basic karyotypes of all the plant species investigated Nicotiana tomentosiformis, N. kawakamii, N. tomentosa, N. otophora, N. setchellii, N. glutinosa (all section Tomentosae), and N. tabacum (tobacco, section Genuinae) are similar (x=12) but the distribution of genic and non-genic repeats is quite variable, making the karyotypes distinct. We found sequence dispersal, and locus gain, amplification and loss, all within the regular framework of the basic genomic structure. We predict that the GRD classes of sequence integrated into an ancestral genome only once in the evolution of section Tomentosae and thereafter spread by vertical transmission and speciation into four species. Since GRD is similar to a transgenic construct that was inserted into the N. tabacum genome, its fate over evolutionary time is interesting in the context of the debate on genetically modified organisms and the escape of genes into the wild. Nicotiana tabacum is thought to be an allotetraploid between presumed progenitors of N. sylvestris (maternal, S-genome donor) and a member of section Tomentosae (T-genome donor). Of section Tomentosae, N. tomentosiformis has the most similar genome to the T genome of tobacco and is therefore the most likely paternal genome donor. It is known for N. tabacum that gene conversion has converted most 18S-5.8S-26S rDNA units of N. sylvestris origin into units of an N. tomentosiformis type. Clearly if such a phenomenon were widespread across the genome, genomic in situ hybridisation (GISH) to distinguish the S and T genomes would probably not work since conversion would tend to homogenise the genomes. The fact that GISH does work suggests a limited role for gene conversion in the evolution of N. tabacum.

Notes: : The efficacy of 2% molecular weight 240, 2% molecular weight 360 polylactic acid (PLA), and an equal mix of both at reducing numbers of Escherichia coli O157:H7 and Lactobacillus plantarum on raw beef was determined. Fresh beef cubes inoculated with either organism were dipped in PLA solutions or wrapped in PLA-sprayed films. Samples were vacuum packaged and stored at 4°C for 42 d. Treated samples maintained a significantly lower pH than controls. Growth of E. coli O157:H7 was totally inhibited by both PLA treatments by up to 7.29 log10 CFU/cm2 when the spray method was used. However, PLA treatments against L. plantarum were not very effective.

Notes: : Glycoprotein of Solanum nigrum Linne (SNL glycoprotein) was isolated and identified using SDS-PAGE. SNL glycoprotein's antioxidative effects were tested in vitro and its cytotoxicity effects were tested using MCF-7 cells. One glycoprotein was isolated from SNL fruits, the other from stems and leaves. SNL glycoprotein is reactive with oxygen radicals, such as the -OH and O2- in vitro. Moreover, the SNL glycoprotein's radical scavenging activity is sensitive to the superoxide anion radical and the hydroxyl radical. In the MCF-7 cell, SNL glycoprotein I had a cytotoxic effect at 1 μg/mL and SNL glycoprotein II at 100 μg/mL. When MCF-7 cells were treated with SNL glycoproteins, nitrite oxide production was induced.

Notes: Growth of gallium oxide on n-GaN was realized in H2O by bias-assisted photoelectrochemical (PEC) oxidation using Al as a counterelectrode instead of a Pt commonly used in the PEC process. Although the growth of the oxide was not observed at below 2 V, the initial oxide growth rate of 8.7 nm/min was shown at a bias of 15 V and ultraviolet light intensity of 300 mW/cm2. However, the growth rate lowered and oxide thickness was saturated to 340 nm. The saturated oxide thickness and initial growth rate were increased with the applied bias. The homogeneous oxide growth and near stoichiometric composition of Ga2O3 were observed in Auger electron spectroscopy analysis results. © 2002 American Institute of Physics.

Notes: Tip characteristics play an important role in the resolution and sensitivity of scanning probe microscopy. Extensive efforts have been devoted to tip fabrication. Most of the research is focused on scanning tunneling microscopy applications, which require sharp and short tips. Long tips that can be bent into cantilevered tips have great potential in atomic force microscopy/apertureless near-field scanning optical microscopy applications. However, the fabrication of such tips has been rarely reported. The present work is carried out with the aim of optimizing the conditions suitable for fabricating long and sharp tungsten tips. Besides topography, optical, and spectroscopic information, electrical and magnetic measurements can also be carried out with such tips obtained with the recipe reported in this article. The long tips also make it possible to measure deep grooves/trenches. © 2002 American Institute of Physics.

Notes: Temperature-dependent Hall effect measurements on unintentionally doped n-type GaN epilayers show that, above room temperature, the Hall-mobility values of different samples vary parallel with each other with temperature. We demonstrate that this anomaly is mainly due to a conductive layer near the GaN/sapphire interface for thin samples with low carrier density. Through trapping electrons, threading edge dislocations (TEDs) debilitate the epilayer contribution in a two-layer mixed conduction model involving the epilayer and the near-interface layer. The trapping may, in part, explain low mobility and anomalous transport in pure GaN layers. Scattering by TEDs is important only at low temperatures. © 2000 American Institute of Physics.

Notes: A two-band model involving the heavy- and light-hole bands was adopted to analyze the temperature-dependent Hall effect measured on Mg-doped p-type GaN epilayers. At 300 K, the hole concentration was determined to be nearly twice the Hall concentration estimated from the measured Hall coefficient, meanwhile the Hall mobility of heavy hole turned out to be only half of the measured one. It is shown that the scattering by space charge and acoustic deformation potential is anomalously enhanced in Mg-doped GaN, and that the light hole affects conspicuously the observed transport parameters. © 2000 American Institute of Physics.

Notes: Cynipid gall formation is achieved by an insect–plant interaction whereby cynipid gallwasps redirect host-plant development to form novel structures to protect and nourish the developing larvae. Work was carried out to investigate the molecular mechanisms involved in this interaction, and extend the understanding of plant tissue development. Cytological changes of the inner-gall tissue throughout the development of several gall species was investigated and the developmental stages of gall formation defined, to reveal two different patterns of development followed by the galls tested. Fluorescent in situ hybridization demonstrated many of the inner-gall cells to be polytenized. Comparisons between inner-gall and non-gall tissue protein signatures by Schönrogge et al. (Plant, Cell and Environment 23, 215–222, 2000) have demonstrated the variation between gall and non-gall protein signatures, and identified a number of inner-gall proteins. Further analysis of one of these inner-gall proteins involved in lipid synthesis, putative biotin carboxyl carrier protein (BCCP), revealed differential expression throughout development, and showed this expression to be concentrated in the inner-gall tissue in all the gall species tested.

Notes: Abstract Cytosine methylation levels and susceptibility to drug-induced hypomethylation have been studied in several Nicotiana tabacum (tobacco) DNA repetitive sequences. It has been shown using HapII, MspI, BamHI and Sau3AI methylation-sensitive restriction enzymes that the degree of 5′-mCmCG-3′ methylation varied significantly between different repeats. There were almost saturation levels of 5-methylcytosine at the inner (3′) cytosine position and variable degrees of methylation at the outer (5′) cytosine at the enzyme recognition sites. The non-transcribed high copy satellite sequences (HRS60, GRS) displayed significant heterogeneity in methylation of their basic units while middle repetitive sequences (R8.1, GRD5, 5S rDNA) were more uniformly modified at both cytosine residues. Dihydroxypropyladenine (DHPA) treatment, which is thought to reduce DNA methyltransferase activity by increasing S-adenosylhomocysteine levels, resulted in extensive demethylation of the outer cytosine in all repeats, and the partial hypomethylation of cytosines at the inner positions in less densely methylated repeats such as HRS60 and GRS. The results suggest that hypomethylation of 5′-mCmCG-3′ sites with DHPA is a gradual non-random process proceeding in the direction mCmCG→CmCG→CCG. The 18S-5.8S-25S rDNA was remarkably hypomethylated relative to the 5S rDNA at all restriction sites studied. Fluorescence in-situ hybridization showed that DNA decondensation within and between the 18S-5.8S-25S and 5S rDNA loci was variable in different nuclei. All nuclei had condensed and decondensed sequence. The chromatin of 18S-5.8S-25S rDNA was more readily digested with micrococcal nuclease than the 5S rDNA suggesting that the overall levels of decondensation were higher for 18S-5.8S-25S rDNA. Variable decondensation patterns within and between loci were also observed for GRS and HRS60. Cytosine methylation of the tobacco repeats is discussed with respect to transcription, overall levels of condensation and overall structure.