High-resolution nonlinear optical imaging of live cells by second harmonic generation

By adapting a laser scanning microscope with a titanium sapphire femtosecond pulsed laser and transmission optics, we are able to produce live cell images based on the nonlinear optical phenomenon of second harmonic generation (SHG). Second harmonic imaging (SHIM) is an ideal method for probing membranes of living cells because it offers the high resolution of nonlinear optical microscopy with the potential for near-total avoidance of photobleaching and phototoxicity. The technique has been implemented on three cell lines labeled with membrane-staining dyes that have large nonlinear optical coefficients. The images can be obtained within physiologically relevant time scales. Both achiral and chiral dyes were used to compare image formation for the case of single- and double-leaflet staining, and it was found that chirality plays a significant role in the mechanism of contrast generation. It is also shown that SHIM is highly sensitive to membrane potential, with a depolarization of 25 mV resulting in an approximately twofold loss of signal intensity.     

Molecular diversity of 1-aminocyclopropane-1-carboxylate (ACC) deaminase producing PGPR from wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) rhizosphere

Aims

The present study was planned to investigate the diversity of 1-aminocyclopropane-1-carboxylate (ACC) deaminase producing bacteria from the rhizosphere of wheat plants and subsequent evaluation of selected PGPR on growth enhancement of wheat seedlings under drought and saline conditions.

Methods

ACC deaminase producing plant growth promoting rhizobacteria (PGPR) were isolated from the rhizosphere of wheat and identified using 16S rRNA gene sequence analysis. Isolates were evaluated for various direct and indirect plant growth promoting (PGP) traits. Plant inoculation experiment was conducted using isolates IG 19 and IG 22 in wheat to assess their plant growth promotion potential under salinity and drought stress.

Results

Thirty-eight ACC deaminase producing PGPR were isolated which belonged to 12 distinct genera and falling into four phyla γ-proteobacteria, β-proteobacteria, Flavobacteria and Firmicutes. Klebsiella sp. was the most abundant genera and followed by Enterobacter sp. The isolates exhibited ACC deaminase activities ranging from 0.106–0.980 μM α- ketobutyrate μg protein^?1 h^?1. The isolates showed multiple PGP traits such as IAA production, phosphate, zinc, potassium solubilization and siderophore production. Enterobacter cloacae (IG 19) and Citrobacter sp. (IG 22) inoculated wheat seedlings showed notable increases in fresh and dry biomass under non-stress as well as under stressed condition.

Conclusion

To the best of our knowledge this is the first report of presence of ACC deaminase activity and other PGP traits from the genus Citrobacter and Empedobacter. Our finding revealed that the γ-proteobacteria group dominated the wheat rhizosphere. Plant inoculation with PGPR could be a sustainable approach to alleviate abiotic stresses in wheat plants. These native PGPR isolates could be used as potential biofertilizers for sustainable agriculture.

     

Production of β-xylosidase from <Emphasis Type="Italic">Trichoderma asperellum</Emphasis> KIF125 and its application in efficient hydrolysis of pretreated rice straw with fungal cellulase

On-site cellulase and hemicellulase production is a promising way to reduce enzyme cost in the commercialization of the lignocellulose-to-ethanol process. A hemicellulase-producing fungal strain suitable for on-site enzyme production was selected from cultures prepared using wet disc-milling rice straw (WDM-RS) and identified as Trichoderma asperellum KIF125. KIF125 hemicellulase showed uniquely high abundance of β-xylosidase in the xylanolytic enzyme system compared to other fungal hemicellulase preparations. Supplementation of Talaromyces cellulolyticus cellulase with KIF125 hemicellulase was more effective than that with the hemicellulases from other fungal sources in reducing the total enzyme loading for the improvement of xylose yield in the hydrolysis of ball-milling RS, due to its high β-xylosidase dominance. β-Xylosidase in KIF125 hemicellulase was purified and classified as a glycosyl hydrolase family 3 enzyme with relatively high specificity for xylobiose. The production of KIF125 β-xylosidase in the fermentor was estimated as 118 U/g-WDM-RS (2350 U/L culture) at 48 h. These results demonstrate that KIF125 is promising as a practical hemicellulase source to combine with on-site cellulase production using T. cellulolyticus.     

外源基因表达造成的群体感应细菌生存压力的建模分析

外源基因的表达及其对细菌种群的影响对于群体感应系统和合成生物学产业的研究具有重要意义。然而,人们对于表达外源蛋白的细菌本身的行为模式仍然知之甚少。为了研究菌落生长和外源基因表达的过程究竟受到哪些因素的影响,文中测量了受Lux类受体调控的外源基因在N-酰基高丝氨酸内酯 (N-acyl homoserine lactone,N-AHL) 信号分子诱导下的表达,并模拟了其对细菌种群动态的影响。文中建立了一个假设性的数学模型,对信号分子诱导表达下细菌种群生长受影响的现象进行了分析。先前的研究通常将细菌种群生长受群体感应系统影响的现象归咎于合成群体感应信号分子的消耗与N-AHL信号分子的毒性,文中提供了对于这种生存压力的另一种可能的解释。     

Complete genome sequence of the bacterium Ketogulonicigenium vulgare Y25

Ketogulonicigenium vulgare is characterized by the efficient production of 2KGA from L-sorbose. Ketogulonicigenium vulgare Y25 is known as a 2-keto-L-gulonic acid-producing strain in the vitamin C industry. Here we report the finished, annotated genome sequence of Ketogulonicigenium vulgare Y25.     

Comparison of three ELISA kits for the differentiation of foot-and-mouth disease virus-infected from vaccinated animals

A study was performed to validate 3 FMDV 3ABC-I-ELISA kits developed in China for the differentiation of FMDV infected and vaccinated animals. Sets of sera from naive and vaccinated cattle as well as from cattle that had been infected were tested for antibodies against nonstructural proteins (NSPs) of FMDV by commercial diagnosis kits, Ceditest® FMDV-NS (Ceditest® kit), UBI® FMDV NONSTRUCTURAL PROTEIN ELISA DIRECTION INSERT (UBI® kit) and a FMDV 3ABC-I-ELISA kit developed at the Lanzhou Veterinary Research Institute. The test parameters (sensitivity and specificity) of the three kits were determined, and the result obtained from FMD 3ABC-I-ELISA kit was compared with that obtained from two foreign kits. The results indicated that the coincidence rate between the FMDV 3ABC-I-ELISA and Ceditest® kits was 98.05%, and the coincidence rate between the FMDV 3ABC-I-ELISA and UBI® kits was 94.4%; the sensitivity of both Ceditest® and FMDV 3ABC-I-ELISA kit was 100%. However, the sensitivity of the UBI® kit was only 81.8%. With sera from naive or vaccinated non-infected animals, the specificity of all tests exceeded 90%.     

A modified Coomassie Brilliant Blue staining method at nanogram sensitivity compatible with proteomic analysis

A more sensitive and convenient Coomassie Brilliant Blue (CBB) staining method for visualizing proteins was developed. Compared with the modifications include the supplement of 10% (v/v) methanol into the fixing solution, an increase of an additional sensitization step and CBB raised from 0.1 to 0.125%. The improved method can detect proteins at nanogram level. The improved method is more sensitive than Blue Silver and more convenient than the Silver protocol. Mass spectrometry results confirmed that it is suitable for subsequent proteomic research.     

Chondroitin sulfates and their binding molecules in the central nervous system

Chondroitin sulfate (CS) is the most abundant glycosaminoglycan (GAG) in the central nervous system (CNS) matrix. Its sulfation and epimerization patterns give rise to different forms of CS, which enables it to interact specifically and with a significant affinity with various signalling molecules in the matrix including growth factors, receptors and guidance molecules. These interactions control numerous biological and pathological processes, during development and in adulthood. In this review, we describe the specific interactions of different families of proteins involved in various physiological and cognitive mechanisms with CSs in CNS matrix. A better understanding of these interactions could promote a development of inhibitors to treat neurodegenerative diseases.     

Winter spatial distribution of fish larvae assemblages relative to the hydrography of the waters surrounding Taiwan

The aim of this study was to investigate the species composition and distribution of fish larvae in relation to hydrographic conditions in the waters surrounding Taiwan Island (TI) in February 2003. In total, 242 kinds of fish larvae belonging to 127 genera and 75 families were recognized. Among these, 109 taxa were identified to the family or genus level, others to the species level. The 12 predominant types, which constituted 71% of the total fish larvae, were Engraulis japonica, Scomber sp., Diaphus spp., Benthosema pterotum, Carangoides ferdau, Embolichthys mitsukurii, Maurolicus sp., unidentified Myctophidae, Gonostoma gracile, Trichiurus lepturus, unidentified Gobiidae, and Myctophum asperum. The distribution of fish larvae showed a clear association with water masses around TI, with higher abundances and lower species richness northwest of TI where the China Coastal Current prevails, and lower abundances and higher species diversity east of TI where the Kuroshio Current dominates. Cluster analysis distinguished three station groups and four species groups, and the distribution patterns of fish larvae also corresponded to hydrographic conditions. The total abundances of fish larvae and eight of the 12 predominant taxa showed significant and positive correlations with zooplankton abundance, which suggests that food source might be a key factor determining the abundance and distribution of fish larvae during the winter.     

3D-QSAR studies of checkpoint kinase 1 inhibitors based on molecular docking and CoMFA

Three-dimensional quantitative structure–activity relationship (3D-QSAR) studies were performed on a series of substituted 1,4-dihydroindeno[1,2-c]pyrazoles inhibitors, using molecular docking and comparative molecular field analysis (CoMFA). The docking results from GOLD 3.0.1 provide a reliable conformational alignment scheme for the 3D-QSAR model. Based on the docking conformations and alignments, highly predictive CoMFA model was built with cross-validated q ² value of 0.534 and non-cross-validated partial least-squares analysis with the optimum components of six showed a conventional r ² value of 0.911. The predictive ability of this model was validated by the testing set with a conventional r ² value of 0.812. Based on the docking and CoMFA, we have identified some key features of the 1,4-dihydroindeno[1,2-c]pyrazoles derivatives that are responsible for checkpoint kinase 1 inhibitory activity. The analyses may be used to design more potent 1,4-dihydroindeno[1,2-c]pyrazoles derivatives and predict their activity prior to synthesis.