Protein Microarray On-Demand: A Novel Protein Microarray System

We describe a novel, simple and low-cost protein microarray strategy wherein the microarrays are generated by printing expression ready plasmid DNAs onto slides that can be converted into protein arrays on-demand. The printed expression plasmids serve dual purposes as they not only direct the synthesis of the protein of interest; they also serve to capture the newly synthesized proteins through a high affinity DNA-protein interaction. To accomplish this we have exploited the high-affinity binding (∼3–7×10 −13 M) of E. coli Tus protein to Ter, a 20 bp DNA sequence involved in the regulation of E. coli DNA replication. In our system, each protein of interest is synthesized as a Tus fusion protein and each expression construct directing the protein synthesis contains embedded Ter DNA sequence. The embedded Ter sequence functions as a capture reagent for the newly synthesized Tus fusion protein. This “all DNA” microarray can be converted to a protein microarray on-demand without need for any additional capture reagent.

Impaired Cognitive Function and Altered Hippocampal Synaptic Plasticity in Mice Lacking Dermatan Sulfotransferase Chst14/D4st1

Chondroitin sulfate (CS) and dermatan sulfate (DS) proteoglycans (PGs) are major extracellular matrix (ECM) components of the central nervous system (CNS). A large body of evidence has shown that CSPGs/DSPGs play critical roles in neuronal growth, axon guidance, and plasticity in the developing and mature CNS. It has been proposed that these PGs exert their function through specific interaction of CS/DS chains with its binding partners in a manner that depends on the sulfation patterns of CS/DS. It has been reported that dermatan 4-O-sulfotransferase-1 (Chst14/D4st1) specific for DS, but not chondroitin 4-O-sulfotransferase-1 (Chst11/C4st1) specific for CS, regulates proliferation and neurogenesis of neural stem cells (NSCs), indicating that CS and DS play distinct roles in the self-renewal and differentiation of NSCs. However, it remains unknown whether specific sulfation profiles of DS has any effect on CNS plasticity. In the present study, Chst14/D4st1-deficient (Chst14−/−) mice was employed to investigate the involvement of DS in synaptic plasticity. First, behavior study using Morris Water Maze (MWM) showed that the spatial learning and memory of Chst14−/− mice was impaired when compared to their wild type (WT) littermates. Corroborating the behavior result, long-term potentiation (LTP) at the hippocampal CA3-CA1 connection was reduced in Chst14−/− mice compared to the WT mice. Finally, the protein levels of N-Methyl-D-aspartate (NMDA) receptor, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor, postsynaptic density 95 (PSD95), growth associated protein 43 (GAP-43), synaptophysin (SYN) and N-ethylmaleimide sensitive factor (NSF) which are important in synaptic plasticity were examined and Chst14/D4st1 deficiency was shown to significantly reduce the expression of these proteins in the hippocampus. Further studies revealed that Akt/mammalian target rapamycin (mTOR) pathway proteins, including protein kinase B (p-Akt), p-mTOR and p-S6, were significantly lower in Chst14−/− mice, which might contribute to the decreased protein expression. Together, this study reveals that specific sulfation of DS is critical in synaptic plasticity of the hippocampus and learning and memory, which might be associated with the changes in the expression of glutamate receptors and other synaptic proteins though Akt/mTOR pathway

The Molecular Signature Underlying the Thymic Migration and Maturation of TCRαβ+CD4+CD8- Thymocytes

BACKGROUND: After positive selection, the newly generated single positive (SP) thymocytes migrate to the thymic medulla, where they undergo negative selection to eliminate autoreactive T cells and functional maturation to acquire immune competence and egress capability. METHODOLOGY/PRINCIPAL FINDINGS: To elucidate the genetic program underlying this process, we analyzed changes in gene expression in four subsets of mouse TCRαβ(+)CD4(+)CD8(-) thymocytes (SP1 to SP4) representative of sequential stages in a previously defined differentiation program. A genetic signature of the migration of thymocytes was thus revealed. CCR7 and PlexinD1 are believed to be important for the medullary positioning of SP thymocytes. Intriguingly, their expression remains at low levels in the newly generated thymocytes, suggesting that the cortex-medulla migration may not occur until the SP2 stage. SP2 and SP3 cells gradually up-regulate transcripts involved in T cell functions and the Foxo1-KLF2-S1P(1) axis, but a number of immune function-associated genes are not highly expressed until cells reach the SP4 stage. Consistent with their critical role in thymic emigration, the expression of S1P(1) and CD62L are much enhanced in SP4 cells. CONCLUSIONS: These results support at the molecular level that single positive thymocytes undergo a differentiation program and further demonstrate that SP4 is the stage at which thymocytes acquire the immunocompetence and the capability of emigration from the thymus

Effect of salinity on growth performance and resistance of the clam Cyclina sinensis against Vibrio parahaemolyticus infection

We examined the growth performance, immune parameters and the susceptibility to Vibrio parahaemolyticus in the clam Cyclina sinensis, which had been reared at different salinity levels of 10‰, 20‰ and 30‰ for 60 days. At the end of the feeding experiment, the biggest shell length and body weight was found in 20‰, followed by 30‰ and 10‰. No significant differences in superoxide dismutase (SOD) activity were observed among the clams held in 10‰, 20‰ and 30‰. Na+/K+ -ATPase (NKA) activity of the clams held in 20‰ and 30‰ were significantly lower than that in group 10‰. The lowest activities of lysozyme (LZM) and glutamic-pyruvic transaminase (GPT) were found in group 10‰. The Integrated Biomarker Response index (IBR) values of the clams had an inverse relationship with salinity: 11.28, 3.40 and 2.85 in 10‰, 20‰ and 30‰, respectively. At the end of the feeding experiment, the clams were infected with V. parahaemolyticus. As time after infection goes on, the survival rate of clams reared in 20‰ was not significantly different with the other two treatments from 24 to 48h after infection. However, from 72 to 120h after infection, it was significantly lower than those reared in 10‰, while it was significantly higher than those reared in 30‰. It is concluded that the clam C. sinensis reared in 10‰ seawater may reduce growth performance and immune ability, whereas increase resistance against V. parahaemolyticus infection

Thermal design optimization and analysis on heating load of rural buildings in northern China

The envelope of rural buildings has been lack of effective and reasonable thermal insulation method and therefore its energy consumption has always been high. In order to address this problem, this paper aims to optimize the thermal design of building envelope. The simulation using DesignBuilder software for modeling and analyzing, using the orthogonal experimental design method to study the effects of external wall, external window and roof on heating load, and optimal thermal insulation scheme was obtained, which was 100mm PUF (external wall), 6mm+12mm+6mm low-E glass (external window) and 100mm PUF (roof). Results revealed that the addition of sunspace can significantly reduce the heating load and thus the selection of window thermal insulation material is very important. Compared with the condition of highest heating load, the energy-efficient rate of optimal scheme reached to 21.4%. The results of this study will serve as the idea for optimal design of rural buildings envelope

Enhanced Power Factor and Ultralow Lattice Thermal Conductivity Induced High Thermoelectric Performance of BiCuTeO/BiCuSeO Superlattice

Based on the first-principles calculations, the electronic structure and transport properties of BiMChO (M=Cu and Ag, Ch=S, Se, and Te) superlattices have been studied. They are all semiconductors with indirect band gaps. The increased band gap and decreased band dispersion near the valence band maximum (VBM) lead to the lowest electrical conductivity and the lowest power factor for p-type BiAgSeO/BiCuSeO. The band gap value of BiCuTeO/BiCuSeO decreases because of the up-shifted Fermi level of BiCuTeO compared with BiCuSeO, which would lead to relatively high electrical conductivity. The converged bands near VBM can produce a large effective mass of density of states (DOS) without explicitly reducing the mobility µ for p-type BiCuTeO/BiCuSeO, which means a relatively large Seebeck coefficient. Therefore, the power factor increases by 15% compared with BiCuSeO. The up-shifted Fermi level leading to the band structure near VBM is dominated by BiCuTeO for the BiCuTeO/BiCuSeO superlattice. The similar crystal structures bring out the converged bands near VBM along the high symmetry points Γ-X and Z-R. Further studies show that BiCuTeO/BiCuSeO possesses the lowest lattice thermal conductivity among all the superlattices. These result in the ZT value of p-type BiCuTeO/BiCuSeO increasing by over 2 times compared with BiCuSeO at 700 K

Cutting Path Planning Technology of Shearer Based on Virtual Reality

With regards to the low degree of digitization, lack of real geological terrain, and low degree of automation in the cutting process of the traditional virtual fully mechanized mining face, we studied the key technologies of virtual operation and cutting path planning of the shearer on the three-dimensional (3D) roof and floor based on the virtual reality engine (Unity3D). Firstly, the virtual 3D coal seam was constructed through the 3D geological coordinate data of the mine. On this basis, the shape function of the scraper conveyor with the adaptive configuration on the floor was constructed to obtain the combined operation of the virtual shearer and the scraper conveyor. The movement of the shearer’s walking and height-adjustment was then, analyzed. A strategy for automatic height-adjustment based on the adjustment of the direction of the drum movement is hence, proposed to control the cutting path of the shearer. Finally, different experimental schemes were simulated in the developed prototype system after which each of the schemes was evaluated using the fuzzy comprehensive evaluation method. The results show that the proposed strategy for trajectory control can improve the accuracy and stability of the shearer’s motion trajectory. In Unity3D, the pre-selected schemes and digital and visual planning of coal production are previewed ahead of time, the whole production process can be mapped synchronously in the production process. It is also obtained that the virtual preview and evaluation of the production process can provide some guidance for actual production

Table3_TRcaller: a novel tool for precise and ultrafast tandem repeat variant genotyping in massively parallel sequencing reads.XLSX

Calling tandem repeat (TR) variants from DNA sequences is of both theoretical and practical significance. Some bioinformatics tools have been developed for detecting or genotyping TRs. However, little study has been done to genotyping TR alleles from long-read sequencing data, and the accuracy of genotyping TR alleles from next-generation sequencing data still needs to be improved. Herein, a novel algorithm is described to retrieve TR regions from sequence alignment, and a software program TRcaller has been developed and integrated into a web portal to call TR alleles from both short- and long-read sequences, both whole genome and targeted sequences generated from multiple sequencing platforms. All TR alleles are genotyped as haplotypes and the robust alleles will be reported, even multiple alleles in a DNA mixture. TRcaller could provide substantially higher accuracy (>99% in 289 human individuals) in detecting TR alleles with magnitudes faster (e.g., ∼2 s for 300x human sequence data) than the mainstream software tools. The web portal preselected 119 TR loci from forensics, genealogy, and disease related TR loci. TRcaller is validated to be scalable in various applications, such as DNA forensics and disease diagnosis, which can be expanded into other fields like breeding programs. Availability: TRcaller is available at https://www.trcaller.com/SignIn.aspx.</p

Table2_TRcaller: a novel tool for precise and ultrafast tandem repeat variant genotyping in massively parallel sequencing reads.XLSX

Calling tandem repeat (TR) variants from DNA sequences is of both theoretical and practical significance. Some bioinformatics tools have been developed for detecting or genotyping TRs. However, little study has been done to genotyping TR alleles from long-read sequencing data, and the accuracy of genotyping TR alleles from next-generation sequencing data still needs to be improved. Herein, a novel algorithm is described to retrieve TR regions from sequence alignment, and a software program TRcaller has been developed and integrated into a web portal to call TR alleles from both short- and long-read sequences, both whole genome and targeted sequences generated from multiple sequencing platforms. All TR alleles are genotyped as haplotypes and the robust alleles will be reported, even multiple alleles in a DNA mixture. TRcaller could provide substantially higher accuracy (>99% in 289 human individuals) in detecting TR alleles with magnitudes faster (e.g., ∼2 s for 300x human sequence data) than the mainstream software tools. The web portal preselected 119 TR loci from forensics, genealogy, and disease related TR loci. TRcaller is validated to be scalable in various applications, such as DNA forensics and disease diagnosis, which can be expanded into other fields like breeding programs. Availability: TRcaller is available at https://www.trcaller.com/SignIn.aspx.</p