A bidirectional gene trap construct suitable for T-DNA and Ds-mediated insertional mutagenesis in rice (Oryza sativa L.)

A construct suitable for genome-wide transfer-DNA (T-DNA) and subsequent transposon-based (Ds) gene trapping has been developed for use in rice (Oryza sativa). This T-DNA/Ds construct contains: Ds terminal sequences immediately inside T-DNA borders for subsequent Ds mobilization; promoterless green fluorescent protein (sgfpS65T) and beta-glucuronidase (uidA) reporter genes, each fused to an intron (from Arabidopsis GPA1 gene) to enable bidirectional gene trapping by T-DNA or Ds; an ampicillin resistance gene (bla) and a bacterial origin of replication (ori) to serve as the plasmid rescue system; an intron-containing hygromycin phosphotransferase gene (hph) as a selectable marker or Ds tracer; and an intron-containing barnase gene in the binary vector backbone (VB) to select against transformants carrying unwanted VB sequences. More than a threefold increase over previously reported reporter gene-based gene trapping efficiencies was observed in primary T-DNA/Ds transformant rice lines, returning an overall reporter gene expression frequency of 23%. Of the plant organs tested, 3.3-7.4% expressed either reporter at varying degrees of organ or tissue specificity. Approximately 70% of the right border (RB) flanking sequence tags (FSTs) retained 1-6 bp of the RB repeat and 30% of the left border (LB) FSTs retained 5-23 bp of the LB repeat. The remaining FSTs carried deletions of 2-84 bp inside the RB or 1-97 bp inside the LB. Transposition of Ds from the original T-DNA was evident in T-DNA/Ds callus lines super-transformed with a transposase gene (Ac) construct, as indicated by gene trap reporter activity and rescue of new FSTs in the resulting double transformant lines.     

O-Acetyl location on cepacian, the principal exopolysaccharide of Burkholderia cepacia complex bacteria

Cepacian is an exopolysaccharide produced by the majority of the isolates belonging to the Burkholderia cepacia complex bacteria, a group of 17 species, some of which infect cystic fibrosis patients, sometime with fatal outcome. The repeating unit of cepacian consists of a backbone having a trisaccharidic repeating unit with three side chains, as reported in the formula below. The exopolysaccharide is also acetylated, carrying from one to three acetyl esters per repeating unit, depending on the strain examined. The consequences of O-acetyl substitution in a polysaccharide are important both for its biological functions and for industrial applications, including the preparation of conjugated vaccines, since O-acetyl groups are important immunogenic determinants. The location of acetyl groups was achieved by NMR spectroscopy and ESI mass spectrometry and revealed that these substituents are scattered in non-stoichiometric ratio on many sugar residues in different positions, a feature which adds to the already unique carbohydrate structure of the polysaccharide.     

The facilitatory influence of anterior cingulate cortex on ON-OFF response of tactile neuron in thalamic ventrobasal nucleus

The structures of limbic system have been found to modulate the auditory, visual and pain afferent signals in the related nuclei of thalamus. One of those structures is anterior cingulate cortex (ACC) that influences nocuous response of the pain-sensitive neurons in the ventropos-tero-lateral nucleus of thalamus. Thus, we inferred that ACC would also modulate tactile information at the thalamic level. To test this assumption, single units were recorded extracellularly from thalamic ventrobasal nucleus (VB). Tactile ON-OFF response and the relationship between different patterns of the responses and the parameters of tactile stimulation were examined. Furthermore, the influence of ACC on the tactile ON-OFF response was studied. ACC stimulation was found to produce a facilitatory effect on the OFF-response of ON-OFF neurons. It lowered the threshold of the off-response of that neuron, and therefore changed the response pattern or enhanced the firing rate of the OFF-response of the neuron. The study on rec     

Mutagenic evaluation of clonazepam using in vitro and in vivo bacterial tests

The mutagenicity of an anti-epileptic drug, clonazepam, and its metabolites was studied by three repair (rec, pol and uvr) assays and reversion (Ames') tests. No detectable induction of mutation could be found in any tests by them per se or via metabolism in rat and mouse.     

Solid-phase classical complement activation by C-reactive protein (CRP) is inhibited by fluid-phase CRP-C1q interaction

C-reactive protein (CRP) interacts with phosphorylcholine (PC), Fcgamma receptors, complement factor C1q and cell nuclear constituents, yet its biological roles are insufficiently understood. The aim was to characterize CRP-induced complement activation by ellipsometry. PC conjugated with keyhole limpet hemocyanin (PC-KLH) was immobilized to cross-linked fibrinogen. A low-CRP serum with different amounts of added CRP was exposed to the PC-surfaces. The total serum protein deposition was quantified and deposition of IgG, C1q, C3c, C4, factor H, and CRP detected with polyclonal antibodies. The binding of serum CRP to PC-KLH dose-dependently triggered activation of the classical pathway. Unexpectedly, the activation was efficiently down-regulated at CRP levels > 150 mg/L. Using radial immunodiffusion, CRP-C1q interaction was observed in serum samples with high CRP concentrations. We propose that the underlying mechanism depends on fluid-phase interaction between C1q and CRP. This might constitute another level of complement regulation, which has implications for systemic lupus erythematosus where CRP is often low despite flare-ups.     

Benzimidazole design,synthesis, and docking to build selective carbonic anhydrase VA inhibitors

The similarity of human carbonic anhydrase (CA) active sites makes it difficult to design selective inhibitors for one or several CA isoforms that are drug targets. Here we synthesize a series of compounds that are based on 5-[2-(benzimidazol-1-yl)acetyl]-2-chloro-benzenesulfonamide (1a) which demonstrated picomolar binding affinity and significant selectivity for CA isoform five A (VA), and explain the structural influence of inhibitor functional groups to the binding affinity and selectivity. A series of chloro-substituted benzenesulfonamides bearing a heterocyclic tail, together with molecular docking, was used to build inhibitors that explore substituent influence on the binding affinity to the CA VA isoform.     

Immunolocalization of H(+)-ATPase and IRT1 enzymes in N(2)-fixing common bean nodules subjected to iron deficiency

The demand for iron in leguminous plants increases during symbiosis, as the metal is utilised for the synthesis of various Fe-containing proteins in both plant and bacteroids. However, the acquisition of this micronutrient is problematic due to its low bioavailability at physiological pH under aerobic conditions. Induction of root Fe(III)-reductase activity is necessary for Fe uptake and can be coupled to the rhizosphere acidification capacity linked to the H⁺-ATPase activity. Fe uptake is related to the expression of a Fe²⁺ transporter (IRT1). In order to verify the possible role of nodules in the acquisition of Fe directly from the soil solution, the localization of H⁺-ATPase and IRT1 was carried out in common bean nodules by immuno-histochemical analysis. The results showed that these proteins were particularly abundant in the central nitrogen-fixing zone of nodules, around the periphery of infected and uninfected cells as well as in the vascular bundle of control nodules. Under Fe deficiency an over-accumulation of H⁺-ATPase and IRT1 proteins was observed especially around the cortex cells of nodules. The results obtained in this study suggest that the increase in these proteins is differentially localized in nodules of Fe-deficient plants when compared to the Fe-sufficient condition and cast new light on the possible involvement of nodules in the direct acquisition of Fe from the nutrient solution.     

Fabrication of VB2/Air Cells for Electrochemical Testing

A technique to investigate the properties and performance of new multi-electron metal/air battery systems is proposed and presented. A method for synthesizing nanoscopic VB₂ is presented as well as step-by-step procedure for applying a zirconium oxide coating to the VB₂ particles for stabilization upon discharge. The process for disassembling existing zinc/air cells is shown, in addition construction of the new working electrode to replace the conventional zinc/air cell anode with a the nanoscopic VB₂ anode. Finally, discharge of the completed VB₂/air battery is reported. We show that using the zinc/air cell as a test bed is useful to provide a consistent configuration to study the performance of the high-energy high capacity nanoscopic VB₂ anode.     

SSRHunter，一个本地化的SSR位点搜索软件的开发

基因组研究的发展,使得利用生物信息学方法在一个相对狭小的基因组区段内开发新的SSR标记成为可能。要实现这一点,首先面临的任务就是潜在的SSR位点的搜索。已有的方法或多或少存在这样或那样的缺陷。文章介绍了利用Visual Basic语言开发本地化的SSR位点搜索软件的尝试。所编制的SSRHunter软件能够较出色地完成这一任务。此外,SSRHunter还提供了自动序列预处理,常规的序列整理和序列变换功能以及方便的结果输出方式。