Suitable transfection methods for single particle tracing in plant suspension cells

Background

Besides classical utilization of wood and paper, lignocellulosic biomass has become increasingly important with regard to biorefinery, biofuel production and novel biomaterials. For these new applications the macromolecular assembly of cell walls is of utmost importance and therefore further insights into the arrangement of the molecules on the nanolevel have to be gained. Cell wall recalcitrance against enzymatic degradation is one of the key issues, since an efficient degradation of lignocellulosic plant material is probably the most crucial step in plant conversion to energy. A limiting factor for in-depth analysis is that high resolution characterization techniques provide structural but hardly chemical information (e.g. Transmission Electron Microscopy (TEM), Atomic Force Microscopy (AFM)), while chemical characterization leads to a disassembly of the cell wall components or does not reach the required nanoscale resolution (Fourier Tranform Infrared Spectroscopy (FT-IR), Raman Spectroscopy).

Results

Here we use for the first time Scanning Near-Field Optical Microscopy (SNOM in reflection mode) on secondary plant cell walls and reveal a segmented circumferential nanostructure. This pattern in the 100 nm range was found in the secondary cell walls of a softwood (spruce), a hardwood (beech) and a grass (bamboo) and is thus concluded to be consistent among various plant species. As the nanostructural pattern is not visible in classical AFM height and phase images it is proven that the contrast is not due to changes in surfaces topography, but due to differences in the molecular structure.

Conclusions

Comparative analysis of model substances of casted cellulose nanocrystals and spin coated lignin indicate, that the SNOM signal is clearly influenced by changes in lignin distribution or composition. Therefore and based on the known interaction of lignin and visible light (e.g. fluorescence and resonance effects), we assume the elucidated nanoscale structure to reflect variations in lignification within the secondary cell wall.     

Long-term mild hypoxia does not reduce thermal tolerance or performance of the freshwater prawn Macrobrachium tenellum

The present study was designed to assess the performance of the freshwater prawn Macrobrachium tenellum in optimal and sub-optimal dissolved oxygen conditions, considering increasing environmental pressures. Thermal tolerance and thermal metabolic scope (TMS) with related integrated biomarker response (IBR) were measured in prawns exposed to normoxia (80% air saturation), mild (40% air saturation) and severe hypoxia (25% air saturation) at three exposure time points (10, 20 and 30 days). Effects of hypoxia on thermal tolerance were not detectable over time; they were perhaps masked by hyperventilation, or by an increase or diversion of haemolymph processes. After 30 days, TMS was 11% higher in mild hypoxia compared with normoxia, while it was 64% lower in severe hypoxia, indicating the loss of aerobic metabolism capacity during the latter. Mild-hypoxia prawns maintained a high IBR over time, supported by antioxidant enzyme activities (mainly superoxide dismutase), which helped avoid the serious oxidative damage (proteins and lipids) seen in severe hypoxia animals, as well as lower acetylcholinesterase activity that indicated failure of communication between the nervous and locomotor systems. Our results documented a high tolerance by M. tenellum to mild-hypoxia events, which should be further tested under seasonal and extreme habitat/tank temperatures.     

Distribution of glyphosate-resistant and glyphosate-susceptible hairy fleabane (Conyza bonariensis) in central California and their phenological development

Glyphosate-resistant (GR) hairy fleabane was first documented in California in 2007. A preliminary study comparing plants from this population to a glyphosate-susceptible (GS) population determined that they developed more rapidly than the GS plants. However, it was not known if this was a general characteristic of all GR populations. Therefore, seeds of hairy fleabane were collected from 122 locations in central California. Plants from each location were grown and tested for resistance using a shikimate enzyme assay. Seeds from 90 locations germinated of which 27 % were determined as GR, 21 % as GS, and 52 % as intermediate/mixed. Plants from five GR and five GS populations, as determined by the assay, were randomly selected for comparisons and were grown in 2010/2011. Time taken by each plant to reach key phenological stages was recorded and converted into growing degree day (GDD) using a base temperature of 4.2 °C. The biomass of each plant was recorded at seed set. The five GR and GS plants had similar phenological development rates and biomass. Therefore, the differences previously identified between the two populations were not present in the additionally tested populations. The phenological development was hence described by a common GDD model with the plants requiring 374, 711, 970, 1289, and 1551 GDD, to reach the rosette, bolting, first bud, first flower, and first seed stages, respectively. This study shows widespread distribution of GR hairy fleabane in central California, provides useful information on biology of this weed for management, and suggests possible label changes to herbicide manufacturers.     

Screening techniques and sources of resistance to foliar diseases caused by major necrotrophic fungi in grain legumes

Summary Necrotrophic pathogens of the cool season food legumes (pea, lentil, chickpea, faba bean and lupin) cause wide spread disease and severe crop losses throughout the world. Environmental conditions play an important role in the development and spread of these diseases. Form of inoculum, inoculum concentration and physiological plant growth stage all affect the degree of infection and the amount of crop loss. Measures to control these diseases have relied on identification of resistant germplasm and development of resistant varieties through screening in the field and in controlled environments. Procedures for screening and scoring germplasm and breeding lines for resistance have lacked uniformity among the various programs worldwide. However, this review highlights the most consistent screening and scoring procedures that are simple to use and provide reliable results. Sources of resistance to the major necrotrophic fungi are summarized for each of the cool season food legumes. Marker-assisted selection is underway for Ascochyta blight of pea, lentil and chickpea, and Phomopsis blight of lupin. Other measures such as fungicidal control and cultural control are also reviewed. The emerging genomic information on the model legume, Medicago truncatula, which has various degrees of genetic synteny with the cool season food legumes, has promise for identification of closely linked markers for resistance genes and possibly for eventual map-based cloning of resistance genes. Durable resistance to the necrotrophic pathogens is a common goal of cool season food legume breeders.     

Comparison of the gelatinization behavior of organic and conventional spelt starches assessed by thermal and rheological analyses

The objective of this study was to compare gelatinization properties and molecular composition of starches extracted from locally grown organic and conventional spelt using thermal, rheological, and SEC analyses, along with Concanavalin A method. Organic and conventional spelt was planted in six replicated plots, and the extracted starch was analyzed for their gelatinization properties. DSC showed that the gelatinization temperature ranged from 56.7 to 68.8 °C with an average peak of 62.4 °C, with no evidence for statistical difference in gelatinization properties between treatments. Rheological behavior variation among samples was more pronounced than that between the two growing conditions. The amylose content ranged from 23.0% to 29.8%. There was no significant difference in the molecular weight of amylose and amylopectin irrespective of the plot locations, although a significant difference was found between the amylopectin molecular weight of organic and conventional spelt starches when analyzed collectively. The organic spelt starch studied may substitute the conventional starch when gelatinization behavior is considered.     

燕麦对冠锈病水平抗性的QTL定位

为了确定与燕麦冠锈病水平抗性基因相关的分子标记，建立了具水平抗病性的品系MN841801-1和感病品种Noble-2杂交的F8群体。该群体由158个重组自交系组成。用112个RFLP标记构建了其连锁图，该图由17个连锁群（A-Q）组成，另有23个位点未连锁，大约覆盖了燕麦基因组的1498.2cM。重组自交系群体的抗病性鉴定分别在温室和田间进行。用病斑面积占总叶面积的百分率（PPA）和病害发展曲线下的面积（AUDPC）作为抗病指标，在田间试验中分别探测到3个燕麦冠锈病水平抗性的QTL，依次命名为PRQ1，PRQ2和PRQ3。这3个QTL分别位于连锁群C，P和N上，总共可解释27%的表现型变量。在温室试验中只探测到PRQ1和PRQ2两个QTL，当用AUDPC和PPA分别作为抗病指标时，这两个QTL可分别解释32%和26%的表现型变量。     

Variation in the levels of the different xylanase inhibitors in grain and flour of 20 French wheat cultivars

A xylanase from Aspergillus aculeatus (glycoside hydrolase family 10), uninhibited by TAXI, and a xylanase from Bacillus subtilis (family 11), uninhibited by XIP-I, were selected to quantify the respective apparent levels of XIP-I and TAXI inhibitors, in flours and grains of 20 wheat varieties. The apparent TAXI amount ranged from 0.05 to 0.19 mg/g in flour (mean: 0.11 mg/g) and from 0.07 to 0.2 mg/g in grain (mean: 0.14 mg/g). The range observed for XIP-I was 0.12–0.6 mg/g in flour (mean 0.32 mg/g), and 0.21–0.56 mg/g in grain (mean: 0.41 mg/g). The inhibition profile of the xylanase from A. aculeatus by a crude inhibitor preparation suggested the presence of an additional component in wheat flour, responsible for an increase in inhibition.