Expression of the hygromycin B phosphotransferase gene confers tolerance to the herbicide glyphosate

Escherichia coli cells and tobacco (cv. Xanthi) plants transformed with the hygromycin B phosphotransferase gene were able to grow in culture medium containing glyphosate at 2.0 mM. The growth of tobacco calli in media containing increasing glyphosate concentrations was measured. The ID₅₀ for glyphosate was 1.70±0.03 mM for hygromycin-B resistant plants, and 0.45±0.02 mM for control plants. Regenerated plants and progeny selected for resistance to hygromycin B were tested for glyphosate tolerance by spraying them with Faena herbicide (formulated glyphosate with surfactant) at a dose equal to 0.24 kg/ha. This was two times the dose required to kill 100 percent of the control plants. Phosphotransferase activity was measured in the extracts of the transformed leaves by the incorporation of ³²P from [^–32P]ATP and it was observed that hygromycin B phosphotransferase was able to recognize the molecule of glyphosate as substrate.Abbreviations (Hyg) Hygromycin - (Km) Kanamycin - (Glp) Glyphosate - (Sarc) Sarcosine - (AMPA) Aminomethylphosphonic acid     

Diversity pattern in Sesamum mutants selected for a semi-arid cropping system

Summary Due to the complex requirements of moisture stress, substantial genetic diversity with a wide array of character combinations and effective simultaneous selection for several variables is necessary for improving the productivity and adaptation of a component crop in order for it to fit into a cropping system under semi-arid tropical conditions. Sesamum indicum L. is grown in Venezuela after rice/sorghum/or maize under such conditions. A mutation breeding program was undertaken using six locally adapted varieties to develop genotypes suitable for the above system. The diversity pattern for nine variables was assessed by multivariate analysis in 301 M₄ progenies. Analysis of the characteristic roots and principal components in three methods of selection, i.e., M₂ bulks (A), individual plant selection throughout (B), and selection in M₃ for single variable (C), revealed differences in the pattern of variation between varieties, selection methods, and varieties x methods interactions. Method B was superior to the others and gave 17 of the 21 best M₅ progenies. Piritu and CF varieties yielded the most productive progenies in M₅ and M₆. Diversity was large and selection was effective for such developmental traits as earliness and synchrony, combined with multiple disease resistance, which could be related to their importance by multivariate analyses. Considerable differences in the variety of character combinations among the high yielding. M₅ progenies of CF and Piritu suggested possible further yield improvement. The superior response of Piritu and CF over other varieties in yield and adaptation was due to major changes in plant type and character associations. Multilocation testing of M₅ generations revealed that the mutant progenies had a 40%–100% yield superiority over the parents; this was combined with earliness, synchrony, and multiple disease resistance, and was confirmed in the M₆ generation grown on a commercial scale. This study showed that multivariate analysis is an effective tool for assessing diversity patterns, choice of appropriate variety, and selection methodology in order to make rapid progress in meeting the complex requirements of semi-arid cropping systems.Project supported by the International Atomic Energy Agency, Vienna, Austria     

Circulating immune complexes and in vitro cell reactivity in paracoccidioidomycosis

The severest forms of paracoccidioidomycosis (Pcm) are associated with impaired cell-mediated immunity, a phenomenon that is reversible with therapy. It has been postulated that plasma factors could be responsible for such immune dysfunction. In this report, circulating immune complexes (CIC) were measured by the Raji cell radioimmunoassay (Raji) and by the¹²⁵I-C1q binding assay (C1q-BA) in sera from 14 patients with either active or inactive forms of Pcm and from 15 healthy controls. The C1q-B A revealed significantly elevated levels of CIC in the sera of all but one of the patients. Four of the 8 active (62%) and 2 of the 6 inactive (33%) patients had CIC levels significantly higher than the controls as determined by the Raji test. Significantly increased levels of CIC were detected only in the active patients by the Raji test. The serum of one of the patients, with a generalized infection and depressed lymphocyte responsiveness, was examined and found to contain a factor which depressed the in vitro proliferation of both homologous and normal lymphocytes. We also found that pre-culture of the patients' lymphocytes before stimulation restored their proliferative capacity, and IC were detectable in the culture supernatants. However, the subsequent addition of the patients' serum to such precultured cells did not reinduce the depression. It is suggested therefore, that the depression of T cell responses observed in Pcm is due to the presence of IC which may interact reversibly with the responding cells and/or activate a suppressor cell population whose activity is diminished by preculture.     

蓖麻蚕两种凝集素在蚕体内的出现与分布

蓖麻蚕Philosamia cynthia ricni血淋巴含两种凝集素,一种凝集兔新鲜红血球,凝血活力被L-鼠李糖和D-半乳糖抑制;另一种凝集戊二醛固定的人和鸡的红血球,凝血活力被岩藻糖抑制.它们在蚕的不同生长阶段及在蚕体各组织中的分布和凝血活力显著不同.血淋巴中这两种凝集素的凝血活力明显比其他组织中高.卵中测不到这两种凝集素活力.本文对这两种凝集素在蚕体中可能的生理功能进行了讨论.     

Influence of form of activated charcoal on embryogenic callus formation in coconut (<Emphasis Type="Italic">Cocos nucifera</Emphasis>)

Development of micropropagation protocols for Cocos nucifera has progressed slowly. Activated charcoal is included in the culture medium of each protocol, mainly to prevent tissue browning. Charcoal production procedures can affect the properties of different brands. In this study, eight types of activated charcoal were evaluated for their effects on free 2,4-dichlorophenoxyacetic acid level, pH, conductivity, and osmolarity of the culture medium and on the frequency of embryogenic callus induction. Moreover, the effect of particle size of the optimum charcoal type on embryogenic callus development was also studied. Charcoal type had a significant effect on (Y3) culture medium properties. Free 2,4-D was highest in Reactivos y Productos Químicos Finos-containing medium and pH was lowest in MERCK-containing medium. Charcoal type also influenced embryogenic callus induction, with acid washed for plant cell and tissue culture-, DARCO- and United States Pharmacopeia-containing media promoting ~60% embryogenic callus, but with different optimal 2,4-D concentrations. Particle size profiles varied among all charcoal types, although small particle fraction (<38 μm) was abundant in all. Use of small particle fractions produced higher frequencies of embryogenic callus (70%) than either large particle or whole charcoal fractions.     

The cysteine 354 and 277 residues of Salmonella enterica serovar Typhi EnvZ are determinants of autophosphorylation and OmpR phosphorylation

An initial biochemical characterization of the Salmonella enterica serovar Typhi ( S . Typhi) EnvZ sensor protein and several mutant derivatives was performed. Autophosphorylation levels were higher for Escherichia coli EnvZ, intermediate for S. enterica serovar Typhimurium EnvZ and very low for S . Typhi EnvZ, in spite of their high amino acid sequence identity. Consequently, OmpR phosphorylation was related to EnvZ autophosphorylation. Among the mutant derivatives, a C354G mutation in S . Typhi EnvZ resulted in a substantial increase in autophosphorylation, while mutation of its other cysteine residue at position 277 to L or S decreased the EnvZ autophosphorylation level. Upon heterodimerization, the S . Typhi C354G mutant complemented the wild type in vitro , increasing the EnvZ-P yield of both monomers, in accordance with the model where EnvZ autophosphorylation occurs in trans , indicating that dimer formation is a dynamic process. Hence, the C354 and the C277 residues are fundamental in determining the particular intrinsic biochemical characteristics of EnvZ.     

HistoChoice® as an alternative to formalin fixation of undecalcified bone specimens

We compared histochemical and immunohistochemical staining as well as fluorochrome labeling in murine bone specimens that were fixed with 10% neutral buffered formalin to those fixed with HistoChoice®. We showed that sections from undecalcified tibiae fixed for 4 h in HistoChoice® resulted in enhanced toluidine blue and Von Kossa histochemical staining compared to formalin fixation. HistoChoice® produced comparable or improved staining for alkaline phosphatase. Acid phosphatase localization was better in formalin fixed specimens, but osteoclasts were visuralized more easily in HistoChoice® fixed specimens. As expected, immunohistochemical labeling was antibody dependent; some antibodies labeled better in HistoChoice® fixed specimens while others were better in formalin fixed specimens. Toluidine blue, Von Kossa, and alkaline phosphatase staining of sections fixed for 12 h produced sections that were similar to 4 h fixed sections. Fixation for 12 h preserved acid phosphatase activity better. Increasing fixation to 12 h affected immunolocalization differentially. Bone sialoprotein labeling in HistoChoice® fixed specimens was comparable to formalin fixed samples. On the other hand, after 12 h formalin fixation, osteocalcin labeling was comparable to HistoChoice®. For most histochemical applications, fixing murine bone specimens for 4 h with HistoChoice® yielded superior staining compared to formalin fixation. If immunohistochemical localization is desired, however, individual antibodies must be tested to determine which fixation process retains antigenicity better. In addition, there was no detectable difference in the intensity of fluorochrome labeling using either fixative. Finally, fixation duration did not alter the intensity of labeling.