Effects of Elevated Carbon Dioxide on the Growth and Foliar Chemistry of Transgenic Bt Cotton

A field study was carried out to quantify plant growth and the foliar chemistry of transgenic Bacillus thuringiensis （Bt） cotton （cv. GK-12） exposed to ambient CO2 and elevated （double-ambient） CO2 for different lengths of time （1, 2 and 3 months） in 2004 and 2005. The results indicated that CO2 levels significantly affected plant height, leaf area per plant and leaf chemistry of transgenic Bt cotton. Significantly, higher plant height and leaf area per plant were observed after cotton plants that were grown in elevated CO2 were compared with plants grown in ambient CO2 for 1, 2 and 3 months in the investigation. Simultaneously, significant interaction between CO2 level x investigating year was observed in leaf area per plant. Moreover, foliar total amino acids were increased by 14%, 13%, 11% and 12%, 14%, 10% in transgenic Bt cotton after exposed to elevated CO2 for 1, 2 or 3 months compared with ambient CO2 in 2004 and 2005, respectively. Condensed tannin occurrence increased by 17%, 11%, 9% in 2004 and 12%, 11%, 9% in 2005 in transgenic Bt cotton after being exposed to elevated CO2 for 1, 2 or 3 months compared with ambient CO2 for the same time. However, Bt toxin decreased by 3.0%, 2.9%, 3.1% and 2.4%, 2.5%, 2.9% in transgenic Bt cotton after exposed to elevated CO2 for 1, 2 or 3months compared with ambient CO2 for same time in 2004 and 2005, respectively. Furthermore, there was prominent interaction on the foliar total amino acids between the CO2 level and the time of cotton plant being exposed to elevated CO2. It is presumed that elevated CO2 can alter the plant growth and hence ultimately the phenotype allocation to foliar chemistical components of transgenic Bt cotton, which may in turn, affect the plant-herbivore interactions.     

Rapid Genetic Analysis of Epithelial-Mesenchymal Signaling During Hair Regeneration

Hair follicle morphogenesis, a complex process requiring interaction between epithelia-derived keratinocytes and the underlying mesenchyme, is an attractive model system to study organ development and tissue-specific signaling. Although hair follicle development is genetically tractable, fast and reproducible analysis of factors essential for this process remains a challenge. Here we describe a procedure to generate targeted overexpression or shRNA-mediated knockdown of factors using lentivirus in a tissue-specific manner. Using a modified version of a hair regeneration model ^{5, 6, 11}, we can achieve robust gain- or loss-of-function analysis in primary mouse keratinocytes or dermal cells to facilitate study of epithelial-mesenchymal signaling pathways that lead to hair follicle morphogenesis. We describe how to isolate fresh primary mouse keratinocytes and dermal cells, which contain dermal papilla cells and their precursors, deliver lentivirus containing either shRNA or cDNA to one of the cell populations, and combine the cells to generate fully formed hair follicles on the backs of nude mice. This approach allows analysis of tissue-specific factors required to generate hair follicles within three weeks and provides a fast and convenient companion to existing genetic models.     

Clonal differences in the response of dark and light reactions of photosynthesis to elevated atmospheric CO2 in poplar

Two hybrid poplar (Populus) clones (i.e., fast growing clone Beauprè and slow growing clone Robusta) were grown for two years from cuttings at close spacings in open top chambers (OTCs) under ambient (AC) and elevated [EC = AC + 350 μmol(CO2) mol-1] CO2 treatments. For clone Beauprè no down-regulation of photosynthesis was observed. Two years of growing under EC resulted in an increase in quantum yield of photosystem 2 (PS2), steady state irradiance saturated rate of net photosynthesis (P _Nmax), chlorophyll (Chl) content, and ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPC) activity for this clone. We suppose that under non-limiting conditions of nitrogen and phosphorus content the response to EC was by building up light-harvesting complexes of PS2 and increasing photochemical efficiency of PS2. Due to a high rate of the primary reactions of photosynthesis and a high RuBPCO activity the end product of the response to EC was an increase in PNmax and a larger saccharides content. The Robusta clone showed a depression in the primary reactions of photosynthesis under EC. We found a decrease in quantum yield of PS2, Chl and phosphorus contents, and in RuBPCO activity. However, an increase in PNmax, saccharides content and Chl a/b ratio was observed. We speculate (1) that the phosphorus deficiency in combination with an increase in CO2 concentrations may lead to a potential damage of the assimilation apparatus of the primary reactions of photosynthesis and to a decrease in photochemical efficiency of PS2; (2) that the primary target of "down-regulation" takes place at PS2 for irradiances above 150 μmol m-2 s-1. This revised version was published online in September 2006 with corrections to the Cover Date.     

Human keratinocytes in culture exhibit no response when exposed to short duration, low amplitude, high frequency (900 MHz) electromagnetic fields in a reverberation chamber

We exposed normal human epidermal keratinocytes to short duration, high frequency, and low amplitude electromagnetic fields, similar to that used by mobile phone technologies. We paid particular attention to the control of the characteristics of the electromagnetic environment generated within a mode stirred reverberation chamber (statistical homogeneity and isotropy of the field and SAR distribution). Two non‐thermal exposure conditions were tested on the epidermal cells: 10‐min exposure with a field amplitude of 8 V/m, and 30 min with 41 V/m. Corresponding specific absorption rates ranged from 2.6 to 73 mW/kg (continuous wave, 900 MHz carrier frequency). We collected RNA from cells subjected to these conditions and used it for a large‐scale microarray screening of over 47000 human genes. Under these conditions, exposure of keratinocytes to the electromagnetic field had little effect; only 20 genes displayed significant modulation. The expression ratios were very small (close to 1.5‐fold change), and none of them were shared by the two tested conditions. Furthermore, those assayed using polymerase chain reaction did not display significant expression modulation (overall mean of the exposed samples: 1.20 ± 0.18). In conclusion, the data presented here show that cultured keratinocytes are not significantly affected by EMF exposure. Bioelectromagnetics 32:302–311, 2011. © 2010 Wiley‐Liss, Inc.     

Cell separation mediated by differential rolling adhesion

Recently, we showed a correlation between the maturity of hematopoietic stem and progenitor cells during development and rolling efficiency on selectins. These findings motivated us to explore a novel separation that exploits differences in selectin-mediated rolling adhesion between populations of cells. We extend the use of a previously developed cell-free system to study the separation of populations of sialyl Lewis x (sLe(x))-coated microspheres designed to roll with different average velocities on L-selectin chimeric substrates under well-defined flow. Results show that a separation that exploits differences in average rolling velocities between cell or microsphere populations is attainable. Excellent recovery and purity values for the slower rolling, or more desirable, populations are obtained and can be estimated from rolling velocity measurements. We also assess the feasibility of a selectin-mediated separation of adult bone marrow cell populations using previously obtained rolling velocity and rolling flux data for CD34+ and CD34- adult bone marrow cells on L-selectin substrates. We believe that a cell separation mediated by differential rolling adhesion can be used to enrich populations of hematopoietic stem and progenitor cells from an adult bone marrow cell preparation and that this method possesses several major advantages over existing antibody-mediated cell-affinity chromatography technologies.     

A simplified method for the cultivation of extreme anaerobic Archaea based on the use of sodium sulfite as reducing agent

The extreme sensitivity of many Archaea to oxygen is a major obstacle for their cultivation in the laboratory and the development of archaeal genetic exchange systems. The technique of Balch and Wolfe (1976) is suitable for the cultivation of anaerobic Archaea but involves time-consuming procedures such as the use of air locks and glove boxes. We describe here a procedure for the cultivation of anaerobic Archaea that is more convenient and faster and allows the preparation of liquid media without the use of an anaerobic chamber. When the reducing agent sodium sulfide (Na₂S) was replaced by sodium sulfite (Na₂SO₃), anaerobic media could be prepared without protection from oxygen outside an anaerobic chamber. Exchange of the headspace of serum bottles by appropriate gases was sufficient to maintain anaerobic conditions in the culture media. Organisms that were unable to utilize sulfite as a source for cellular sulfur were supplemented with hydrogen sulfide. H₂S was simply added to the headspace of serum bottles by a syringe. The use of H₂S as a source for sulfur minimized the precipitation of cations by sulfide. Representatives of 12 genera of anaerobic Archaea studied here were able to grow in media prepared by this procedure. For the extremely oxygen-sensitive organism Methanococcus thermolithotrophicus, we show that plates could be prepared outside an anaerobic chamber when sulfite was used as reducing agent. The application of this method may faciliate the cultivation and handling of extreme anaerobic Archaea considerably. Received: January 4, 2000 / Accepted: April 5, 2000     

Anatomical observations on the nucellar apex of Wellwitschia mirabilis and the chemical composition of the micropylar drop

Summary In the young ovule of Welwitschia mirabilis the nucellar apex is dome shaped and starch begins to accumulate near the female gametophyte. With the degeneration of the cells of the nucellar apex, a pollen chamber is formed, which contains the micropylar fluid. Starch storage increases considerably in the upper part of the nucellus. Pollen drop emission is not a rhythmic process, and pollination does not produce the rapid withdrawal of droplets. The micropylar drop consists almost entirely of sugars, uronic acids and a very small amount of free amino acids and enzymes. The mechanism of micropylar drop secretion and its probable role in the process of pollination is discussed.This work was supported by a grant from MURST 40%     

Survival and recovery of perennial forage grasses under prolonged Mediterranean drought: II. Water status, solute accumulation, abscisic acid concentration and accumulation of dehydrin transcripts in bases of immature leaves

Effects of ozone on spring wheat ( Triticum aestivum L. cv. Satu) were studied in an open-top chamber experiment during two growing seasons (1992–1993) at Jokioinen in south-west Finland. The wheat was exposed to filtered air (CF), non-filtered air (NF), non-filtered air+35 nl l⁻¹ ozone for 8 h d⁻¹ (NF⁺) and ambient air (AA). Each treatment was replicated five times. Two wk after anthesis, after 4 wk of ozone treatment (NF⁺, 45 nl l⁻¹ 1000–1800 hours, seasonal mean) the net CO₂ uptake of wheat flag leaves was decreased by c . 40% relative to CF and NF treatments, both initial and total activity of Rubisco and the quantity of protein-bound SH groups were decreased significantly. Added ozone also significantly accelerated flag leaf senescence recorded as a decrease in chloroplast size. The effect was significant 2 wk after anthesis, and senescence was complete after 4 wk. In the CF and NF treatments senescence was complete 5 wk after anthesis. The significant effect of ozone on the chloroplasts and net CO₂ uptake 2 wk after anthesis did not affect the grain filling rate. However, since the grain filling period was shorter for ozone fumigated plants, kernels were smaller. The decrease in 1000-grain weight explained most of the yield reduction in the plants under NF⁺ treatment. The results indicate that wheat plants are well buffered against substantial decrease in source activity, and that shortened flag leaf duration is the major factor causing ozone-induced yield loss.     

A simple method for screening antimicrobial compounds with application to plant disease and fruit quality

Aims:  It is often difficult to extrapolate information from a Petri dish and apply it to commercial applications as with antimicrobial assays. Often large volumes of commodities are used for a virtually untested protocol and result in an unnecessary expenditure of time and materials. An intermediate method, where experimental compounds could be tested directly on a specific commodity without expending large quantities of either sample or compound, would be practical and economical.
Methods and Results:  A method was developed that employs the use of a small experimental chamber in which pieces of natural materials (e.g. fruit) can be tested with antimicrobial compounds. This method uses a type of autoclavable incubation chamber with a filter paper base, hydrating sponge pieces and a sterile glass sample platform. The chamber offers a sterile, controlled environment and can be manipulated to serve a number of studies.
Conclusion:  The chamber results are more analogous to what happens on whole fruit than the Petri dish.
Significance and Impact of the Study:  This method is presently used to screen for antimicrobial compounds and treatments needed to control serious economic pests compromising Florida's agriculture.