Codon optimization, constitutive expression and antimicrobial characterization of hen egg white lysozyme (HEWL) in Pichia pastoris

Fusarium oxysporum (F. oxysporum) and Verticillium dahlia (V. dahlia) causes severe cotton disease in China and other cotton-producing countries. Hen egg white lysozyme (HEWL) has antimicrobial properties. In this study, a codon-optimized HEWL gene was synthesized and cloned into the yeast expression vector, pPIC9K, under the control of the Pichia pastoris (P. pastoris) glyceraldehyde-3-phosphate dehydrogenase promoter (pGAP). Results showed that codon-optimized HEWL (oHEWL) was constitutively expressed. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) indicated that the molecular weight of recombinant HEWL (roHEWL) was 14 kDa which corresponds to the standard HEWL. The expression of the roHEWL reached to 54 mg/L. Activity of the roHEWL was 1680 U/mL. The optimum pH for roHEWL was from 6.0 to 7.0, and the optimum temperature was 55°C. In vitro antimicrobial activity assay revealed that roHEWL can lyse cell walls of the gram positive bacteria, Micrococcus lysodeikticus (M. lysodeikticus). In vivo studies showed that it inhibits plant fungi, F. oxysporum and V. dahlia. roHEWL anti-fungal properties might be useful for future genetically engineered cotton plant resistance against pathogenic fungal disease.Keywords: Hen egg white lysozyme (HEWL), antimicrobial activity, glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter, codon optimization, constitutive expressio

Experimental Verification of Solid-like and Fluid-like States in the Homogeneous Fluidization Regime of Geldart A Particles

The mechanisms underlying homogeneous fluidization of Geldart A particles have been debated for decades. Some ascribed the stability to interparticle forces, while others insisted a purely hydrodynamic explanation. Valverde et al. (2001) fluidized 8.53-μm (i.e., Geldart C) particles by the addition of fumed silica nanoparticles and found that even during homogeneous fluidization both solid-like and fluid-like behavior can be distinguished. However, it is still unclear whether both states exist for true Geldart A particles. In this paper, the particulate fluidization characteristics of three typical Geldart A powders were studied by camera recording, electrical capacitance tomography, and pressure fluctuation. For the first time, the existence of both a solid-like state dominated by interparticle forces and a fluid-like state dominated by fluid dynamics during homogeneous expansion of Geldart A particles was experimentally verified. Furthermore, the ability and performance of the used measurement techniques to identify different flow regimes were compared.</p

Galectin-1 Promotes Metastasis in Gastric Cancer Through a Sphingosine-1-Phosphate Receptor 1-Dependent Mechanism

Background/Aims: Increased expression of galectin-1 (Gal-1) in gastric cancer (GC) promotes metastasis and correlates with poor prognosis. The mechanisms by which Gal-1 promotes GC metastasis remain unknown. Methods: Gal-1and Sphingosine-1-phosphate receptor 1 (S1PR1) were determined by immunohistochemistry(IHC) and quantitative real time polymerase chain reaction (qRT-PCR) in GC specimens. Stably transfected Gal-1 or S1PR1 into SGC7901 and MGC-803 cells, western blot and invasion assays in vitro and nude mice tumorigenicity in vivo were also employed. Results: Overexpression of Gal-1 enhanced expression of S1PR1 in SGC-7901 cells, and increased cell invasion, while knockdown Gal-1 in MGC-803 cells reduced S1PR1 expression and diminished invasion. Simultaneous knockdown of Gal-1 and overexpression of S1PR1 in MGC803 cells rescued invasive ability of MGC803 cells. S1PR1 was associated with expression of epithelial-to-mesenchymal transition (EMT) markers in vitro and in clinical samples. EMT induced in MGC-803 cells by TGF-β1 was accompanied by S1PR1 activation, while knockdown of S1PR1 reduced response to TGF-β1, suggest that Gal-1 promotes GC invasion by activating EMT through a S1PR1-dependent mechanism. Overexpression of S1PR1 promoted subcutaneous xenograft growth and pulmonary metastases, and enhanced expression of EMT markers. Conclusion: Galectin-1 promotes metastasis in gastric cancer through a S1PR1- dependent mechanism, our results indicate that targeting S1PR1 may be a novel strategy to treat GC metastasis

Study on the Mechanical Performance of H-Shaped Steel-Concrete Laminated Plate Composite Beams under Negative Bending Moment

To make the construction of assembled steel-reinforced truss concrete laminated plate composite structure faster, safer, and more efficient, this paper proposes an H-shaped steel-reinforced truss concrete laminated plate composite structure with new angle connectors embedded in the precast bottom panel. Through experimental studies on the H-shaped steel-concrete laminated plate composite beams with precast bottom panels protruding from the bent-up bars, precast bottom panels with embedded new angle connectors and laminated whole cast slab, the similarities and differences of load-deflection, deflection distribution, interface slip, crack distribution and cross-section strain distribution of three groups of composite beams under negative bending moment were analyzed and compared. Using ABAQUS finite element software, we established a finite element model and found the numerical simulation results were in good agreement with the experimental results. Based on this, five groups of finite element models were established for parametric analysis to investigate the effect of concrete strength on the flexural load capacity and flexural stiffness of the steel-laminated plate composite beams with embedded angle connectors. The results of the study show that the combined performance of the H-shaped steel-concrete laminated plate composite beams with the new angle connection embedded in the precast bottom panel was better and the flexural stiffness was greater. The slippage of the H-shaped steel-concrete laminated plate composite beams with embedded new angle connectors in the precast bottom panel was less than the slippage of the precast bottom slab bent-up bars protruding and the laminated cast plate, with the maximum slippage being only 1/2 of the precast bottom panel bent-up bars protruding. In the composite structure of H-shaped steel-concrete composite slabs under negative bending moment, shear angle connectors can replace the bent-up bars protruding from the laminated bottom panel to achieve without extending the reinforcement of the laminated bottom panel

Study on the Mechanical Performance of H-Shaped Steel-Concrete Laminated Plate Composite Beams under Negative Bending Moment

To make the construction of assembled steel-reinforced truss concrete laminated plate composite structure faster, safer, and more efficient, this paper proposes an H-shaped steel-reinforced truss concrete laminated plate composite structure with new angle connectors embedded in the precast bottom panel. Through experimental studies on the H-shaped steel-concrete laminated plate composite beams with precast bottom panels protruding from the bent-up bars, precast bottom panels with embedded new angle connectors and laminated whole cast slab, the similarities and differences of load-deflection, deflection distribution, interface slip, crack distribution and cross-section strain distribution of three groups of composite beams under negative bending moment were analyzed and compared. Using ABAQUS finite element software, we established a finite element model and found the numerical simulation results were in good agreement with the experimental results. Based on this, five groups of finite element models were established for parametric analysis to investigate the effect of concrete strength on the flexural load capacity and flexural stiffness of the steel-laminated plate composite beams with embedded angle connectors. The results of the study show that the combined performance of the H-shaped steel-concrete laminated plate composite beams with the new angle connection embedded in the precast bottom panel was better and the flexural stiffness was greater. The slippage of the H-shaped steel-concrete laminated plate composite beams with embedded new angle connectors in the precast bottom panel was less than the slippage of the precast bottom slab bent-up bars protruding and the laminated cast plate, with the maximum slippage being only 1/2 of the precast bottom panel bent-up bars protruding. In the composite structure of H-shaped steel-concrete composite slabs under negative bending moment, shear angle connectors can replace the bent-up bars protruding from the laminated bottom panel to achieve without extending the reinforcement of the laminated bottom panel

Seismic Performance of a New Assembled Bolt-Connected Concrete Beam–Column Joint: Experimental Test and Finite Element Modeling

A new assembled bolt-connected concrete beam–column joint is proposed, aimed at completing the repair of a post-earthquake node by replacing the bolts and precast beams. Low-cycle loading tests were performed on two new full-scale connections to investigate the effect of bolt strength on the seismic performance of the new connections. A finite element model was established based on the experimental node specimens and compared with the experimental results to verify the accuracy of the finite element simulation results. The seismic performance of the new joints under different axial ratios was studied using finite element software to determine the effect of the axial pressure ratio on the seismic performance of the new joints. Based on the research carried out, a new improved joint was designed, numerical models of the improved joint were established using finite element software, and the seismic performance of the improved joint was compared with the results of the experimental simulation to analyze the seismic performance of the improved joints. The results of the study showed that the bolts and precast concrete beams are the main load-bearing members in the period of service. The joint can be repaired by replacing the bolts and precast concrete beams under seismic action, which meets the new joint design concepts. The finite element simulation results are in good agreement with the experimental results. The larger the axial compression ratio, the earlier the failure stage of the concrete, and the faster the bearing capacity and ductility decrease. The larger the axial compression ratio, the higher the initial stiffness of the joints and the greater the rate of stiffness reduction. The bolt stress distribution of the modified and optimized joints is more satisfactory. This change in node form can improve the recovery efficiency of the joint to a certain extent

Experimental study of tendon sheath repair via decellularized amnion to prevent tendon adhesion.

The adhesion of tendon and surrounding tissue is the most common complication after repairing an injured tendon. The injured flexor tendons in zone II are frequently accompanied by tendon sheath defects, which lead to poor recovery. A variety of biological and non-biological materials have been recently used for repair or as substitute for tendon sheaths to prevent tendon adhesion. However, non-biological materials, such as polyethylene films, have been used to prevent tendon adhesions by mechanical isolation. The possibility of tendon necrosis and permanent foreign body remains due to the lack of permeability and the obstruction of nutrient infiltration. The natural macromolecule amniotic membrane derived from organisms is a semi-permeable membrane with the following characteristics: smooth; without vascular, nerve, and lymphatic; and rich in matrix, cytokines, enzymes, and other active ingredients. The unique structure of this membrane makes it an ideal biomaterial. In the experiment in Henry chicken, the model of tendon sheath defect and the flexor digitorum tendon in zone II was established and randomly divided into control group, medical membrane group, and decellularized amniotic membrane group. Samples were obtained at the 2nd, 4th, 8th, and 12th week after operation. General, histological, and biomechanical tests were performed to investigate the preventive effect of repaired tendon sheath by decallularized amniotic membrane. Experimental results showed the following: the amniotic membrane group and the medical membrane group had mild inflammatory reaction and tissue edema, and nearly no adhesion was observed in the surrounding tissue; the fibroblast-like cells were distributed in layers under the light microscope; the amniotic membrane group was denser than the medical membrane group cells, and numerous fibroblasts were disorganized in the control group. Biomechanical measurements showed that the sliding distance of tendon, the total flexion angle of the toes, and the tendon maximum tensile breaking strength at the early postoperative were significantly better than in the control group. Through this experiment, the amniotic membrane, as a natural biological substitute material in the construction of tendon sheath, can effectively inhibit exogenous healing and promote endogenous healing to prevent tendon adhesion

Magnetic semiconductor in rare-earth-element–based quaternary Heusler compounds

Magnetic semiconductors have become a widely studied direction in spintronic materials due to their unique energy band structure and two different spin transport channels. In this work, we imported rare-earth atoms into quaternary Heusler compounds and considered on Slater-Pauling rules. Magnetic semiconductors FeCrYSn, FeCrYGe, FeCrLuGe, FeCrLuSn, FeCrLuSi, FeVLuSb, MnCrYSb, and MnCrLuSb were found through first-principle calculations. According to the result, the band gaps of these quaternary Heusler compounds are extremely narrow

Segmented transmission delay based decoupling for parallel simulation of a distribution network

Abstract The introduction of inverters and power electronic devices increases the amount of calculation needed for electromagnetic transient simulation in distribution networks with a high proportion of distributed generation. Hence, the parallel calculation of electromagnetic transient simulation has become an effective way to improve its efficiency. To realize efficient parallel simulation, a method to decouple transmission lines in the distribution network based on the segmented transmission delay model is proposed, which divides the line into two segments according to different transmission delays. The transmission delay of one segment is an integral multiple of the simulation step used for line decoupling, whereas the other segment is a lumped parameter model, which makes it equivalent to the original line. In addition, an interface for the distribution network based on the segmented transmission delay model is designed and implemented, and the decoupling simulation of a distribution network is performed. Finally, using the simulation examples of IEEE 34 and IEEE 123 node test feeders, the proposed decoupling method is compared with other established methods. It is found to be more accurate and efficient as well as more suitable for the parallel computation of a distribution network simulation than the comparison methods

Co-expression of nitrogenase proteins in cotton (Gossypium hirsutum L.)

Chemical nitrogen fertilizer can maintain crop productivity, but overuse of chemical nitrogen fertilizers leads to economic costs and environmental pollution. One approach to reduce use of nitrogen fertilizers is to transfer nitrogenase biosynthetic pathway to non-legume plants. Fe protein encoded by nifH and MoFe protein encoded by nifD and nifK are two structural components of nitrogenase. NifB encoded by nifB is a critical maturase that catalyzes the first committed step in the biosynthesis of nitrogenase FeMo-cofactor that binds and reduces N2. Expression of the nifB, nifH, nifD and nifK is essential to generate plants that are able to fix atmospheric N2. In this study, the four genes (nifB, nifH, nifD and nifK) from Paenibacillu polymyxaWLY78 were assembled in plant expression vector pCAMBIA1301 via Cre/LoxP recombination system, yielding the recombinant expression vector pCAMBIA1301-nifBHDK. Then, the four nif genes carried in the expression vector were co-introduced into upland cotton R15 using Agrobacterium tumefaciens-mediated transformation. Homozygous transgenic cotton lines B2, B5 and B17 of T3 generation were selected by PCR and RT-PCR. qRT-PCR showed that nifB, nifH, nifD and nifK were co-expressed in the transgenic cottons at similar levels. Western blotting analysis demonstrated that NifB, NifH, NifD and NifK were co-produced in the transgenic cottons. Co-expression of the four critical Nif proteins (NifB, NifH, NifD and NifK) in cottons represents an important step in engineering nitrogenase biosynthetic pathway to non-legume plants