Simple equations to estimate light interception by isolated trees from canopy structure features: assessment with three-dimensional digitized apple trees

* Simple models of light interception are useful to identify the key structural parameters involved in light capture. We developed such models for isolated trees and tested them with virtual experiments. Light interception was decomposed into the projection of the crown envelope and the crown porosity. The latter was related to tree structure parameters. * Virtual experiments were conducted with three-dimensional (3-D) digitized apple trees grown in Lebanon and Switzerland, with different cultivars and training. The digitized trees allowed actual values of canopy structure (total leaf area, crown volume, foliage inclination angle, variance of leaf area density) and light interception properties (projected leaf area, silhouette to total area ratio, porosity, dispersion parameters) to be computed, and relationships between structure and interception variables to be derived. * The projected envelope area was related to crown volume with a power function of exponent 2/3. Crown porosity was a negative exponential function of mean optical density, that is, the ratio between total leaf area and the projected envelope area. The leaf dispersion parameter was a negative linear function of the relative variance of leaf area density in the crown volume. * The resulting models were expressed as two single equations. After calibration, model outputs were very close to values computed from the 3-D digitized databases.     

Cold glycerol-saline: the promising quenching solution for accurate intracellular metabolite analysis of microbial cells

Microbial metabolomics has been seriously limited by our inability to perform a reliable separation of intra- and extracellular metabolites with efficient quenching of cell metabolism. Microbial cells are sensitive to most (if not all) quenching agents developed to date, resulting in leakage of intracellular metabolites to the extracellular medium during quenching. Therefore, as yet we are unable to obtain an accurate concentration of intracellular metabolites from microbial cell cultures. However, knowledge of the in vivo concentrations of intermediary metabolites is of fundamental importance for the characterization of microbial metabolism so as to integrate meaningful metabolomics data with other levels of functional genomics analysis. In this article, we report a novel and robust quenching method for microbial cell cultures based on cold glycerol-saline solution as the quenching agent that prevents significant leakage of intracellular metabolites and, therefore, permits more accurate measurement of intracellular metabolite concentrations in microbial cells.     

Detection of Salmonella sp. in Dermanyssus gallinae using an FTA filter-based polymerase chain reaction

Salmonella spp. bacteria are responsible for some of the most important zoonoses worldwide. Because Dermanyssus gallinae (DeGeer) (Acari: Dermanyssidae) has been recently reported to be an experimental vector of Salmonella Enteritidis, it would be of benefit to evaluate the presence of this bacterium in mites. A molecular detection tool associating a simple filter-based DNA preparation with a specific 16S rDNA Salmonella sp. polymerase chain reaction (PCR) amplification was described. The limit of detection with this method was 2 x 10(4) bacteria per mite. To adapt this technique for large-scale studies, two sizes of mite pools were tested and a preliminary investigation was carried out on mites from 16 currently or previously contaminated farms. Mites sampled from one farm of each type were positive for Salmonella, suggesting that Dermanyssus could act as a reservoir between flocks. In further investigations, it will be necessary to carry out a large-scale study to assess the role of D. gallinae in the epidemiology of avian salmonellosis.     

Chemical rescue of a site-modified ligand to a [4Fe-4S] cluster in PsaC, a bacterial-like dicluster ferredoxin bound to Photosystem I

Chemical rescue of site-modified amino acids using externally supplied organic molecules represents a powerful method to investigate structure-function relationships in proteins. Here we provide definitive evidence that aryl and alkyl thiolates, reagents typically used for in vitro iron-sulfur cluster reconstitutions, serve as rescue ligands to a site-specifically modified [4Fe-4S]^1+,2+ cluster in PsaC, a bacterial dicluster ferredoxin-like subunit of Photosystem I. PsaC binds two low-potential [4Fe-4S]^1+,2+ clusters termed F_A and F_B. In the C13G/C33S variant of PsaC, glycine has replaced cysteine at position 13 creating a protein that is missing one of the ligating amino acids to iron-sulfur cluster F_B. Using a variety of analytical techniques, including non-heme iron and acid-labile sulfur assays, and EPR, resonance Raman, and Mössbauer spectroscopies, we showed that the C13G/C33S variant of PsaC binds two [4Fe-4S]^1+,2+ clusters, despite the absence of one of the biological ligands. ¹⁹F NMR spectroscopy indicated that the external thiolate replaces cysteine 13 as a substitute ligand to the F_B cluster. The finding that site-modified [4Fe-4S]^1+,2+ clusters can be chemically rescued with external thiolates opens new opportunities for modulating their properties in proteins. In particular, it provides a mechanism to attach an additional electron transfer cofactor to the protein via a bound, external ligand.     

中国白兔白介素-10基因的克隆、表达及其抗体的制备

目的克隆并表达中国白兔IL-10基因,制备抗IL-10多克隆抗体。方法运用RT-PCR对从ConA诱导后的中国白兔外周血单核细胞(PMBCr)总RNA中扩增IL-10基因,克隆后进行测序和遗传进化分析,同时将其亚克隆pET28a中,在大肠杆菌中诱导表达,并用纯化的表达产物制备多克隆抗体。结果该基因全长537 bp,编码178个氨基酸。与欧洲兔IL-10基因同源性很高,但与鼠、鸡、河豚和斑马鱼等不同物种IL-10基因差异较大。经IPTG诱导后,重组菌体裂解物经SDS-PAGE电泳可检测到相对分子质量为23.4×103的重组蛋白。Western blot分析表明,中国白兔IL-10基因已经表达。重组表达的蛋白量可占菌体蛋白的15.2%。结论成功实现了中国白兔IL-10的原核表达,制备了小鼠抗兔IL-10的多克隆抗体。     

用于哺乳动物细胞转染的高纯度质粒DNA的制备

目的：建立简便高效、成本低廉和安全无污染的高纯度质粒提取方法。方法：在乙酸铵方法的基础上加以改进,主要改进之处在于增加了用聚乙二醇纯化质粒的步骤,并对溶液Ⅱ和溶液Ⅲ的成分和具体实验参数也做了合理的改进,以最少的步骤,充分去除了残存杂质,保证了质粒的超纯状态。结果：用本方法提取的质粒与用QIAGEN plasmid midi Kit提取的质粒在理化指标上没有差别,对哺乳动物胞具有同样的转染效率。结论：本方法可完全取代QIAGEN公司的试剂盒用于提取超纯质粒。     

A microaliquoting technique for precise histological annotation and optimization of cell content in frozen tissue specimens

Knowledge of the exact cell content of frozen tissue samples is of growing importance in genomic research. We developed a microaliquoting technique to measure and optimize the cell composition of frozen tumor specimens for molecular studies. Frozen samples of 31 mesothelioma cases were cut in alternating thin and thick sections. Thin sections were stained and evaluated visually. Thick sections, i.e., microaliquots, were annotated using bordering stained sections. A range of cellular heterogeneity was observed among and within samples. Precise annotation of samples was obtained by integration and compared to conventional single face and “front and back” section estimates of cell content. Front and back estimates were more highly correlated with block annotation by microaliquoting than were single face estimates. Both methods yielded discrepant estimates, however, and for some studies may not adequately account for the heterogeneity of mesothelioma or other malignancies with variable cellular composition. High yield and quality RNA was extracted from precision annotated, tumor-enriched subsamples prepared by combining individual microaliquots with the highest tumor cellularity estimates. Microaliquoting provides accurate cell content annotation and permits genomic analysis of enriched subpopulations of cells without fixation or amplification.     

An improved method for the analysis of Cryptosporidium parvum oocysts by matrix-assisted laser desorption/ionization time of flight mass spectrometry

Cryptosporidium parvum oocysts were analyzed using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). Sample preparation proved to be a crucial step in the acquisition of acceptable mass spectra. Oocysts of C. parvum and the matrix were mixed and held for at least 45 min to produce reproducible, representative mass spectra. Sporozoites were also excysted from oocysts, purified, and analyzed using MALDI-TOF MS. The mass spectra of the intact oocysts contained many of the same peaks found in the mass spectra of the sporozoites, suggesting that during analysis, the internal constituents, not just the oocyst wall, are ablated by the laser.