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RNA nanotechnology is a term that refers to the design, fabrication, and utilization of nanoparticles mainly composed of ribonucleic acids via bottom-up self-assembly. The packaging RNA (pRNA) of the bacteriophage phi29 DNA packaging motor has been developed into a nano-delivery platform. This protocol describes the synthesis, assembly, and functionalization of pRNA nanoparticles based on three ‘toolkits’ derived from pRNA structural features: interlocking loops for hand-in-hand interactions, palindrome sequences for foot-to-foot interactions, and an RNA three-way junction for branch-extension. siRNAs, ribozymes, aptamers, chemical ligands, fluorophores, and other functionalities can also be fused to the pRNA prior to the assembly of the nanoparticles, so as to ensure the production of homogeneous nanoparticles and the retention of appropriate folding and function of the incorporated modules. The resulting self-assembled multivalent pRNA nanoparticles are thermodynamically and chemically stable, and they remain intact at ultra-low concentrations. Gene silencing effects are progressively enhanced with increasing number of siRNA in each pRNA nanoparticle. Systemic injection of the pRNA nanoparticles into xenograft-bearing mice has revealed strong binding to tumors without accumulation in vital organs or tissues. The pRNA-based nano-delivery scaffold paves a new way towards nanotechnological application of pRNA-based nanoparticles for disease detection and treatment. The time required for completing one round of this protocol is 3–4 weeks, including in vitro functional assays, or 2–3 months including in vivo studies.

Investigation of an organic extract of the fruits Schisandra wilsoniana led to the isolation of two new highly oxygenated. nortriterpenoids, named schilancidilactones V-W (1-2). Their structures were elucidated by spectroscopic evidence. Compounds 1-2 feature a double bond between C-7 and C-8 compared with related known nortriterpenoids isolated from the genus Schisandra. Compounds 1 and 2 were tested for their anti-HIV-1 activities and cytotoxicity. The results revealed that compounds 1 and 2 showed moderate anti-HIV-1 activities with EC50 3.05 and 2.87 mu g/mL, respectively, and compound 1 showed high cytotoxicity against KB and MDA-MB-231 cell with IC50 values of 3.18 and 5.22 mu M, respectively.

Abscisic acid (ABA) plays important roles in many aspects of plant growth and development. Botrytis cinerea TB-3-H8, a high-yield strain of ABA, was used to elucidate the molecular mechanisms of ABA production in the present work. cDNA-amplified fragment length polymorphism (cDNA-AFLP) technique was applied to isolate genes differentially expressed between ABA high and low-yield conditions. This resulted in the identification of 856 differentially expressed transcript-derived fragments (TDFs). Forty-five TDFs that displayed obvious up-regulated expression profiles in the ABA high-yield condition were sequenced. Based on BlastX in NCBI, 31 TDFs were assumed to have homology with genes encoding proteins with known functions. According to molecular function of gene ontology (GO) analysis, the 31 TDFs were categorized to proteins with enzyme catalytic activities, transcription factor activities, transporter activities, and kinds of binding activities. Further confirmation of the differential expression of these sequences was carried out by performing semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) on 10 randomly selected TDFs. Five up-regulated genes were selected to analyze the expression profiles using real-time PCR. This study enriches our knowledge of the molecular basis for ABA biosynthesis in B. cinerea TB-3-H8.

If tetragonal FePt nanoparticles (NPs) are to be used as magnetic recording media, a means of transforming the particles into the tetragonal phase while maintaining the small particle size must be found. In this work, uniform and high coercivity L10–FePtAg nanoparticles are successfully synthesized by a one-pot polyol-process method, using an octadecylamine solvent. Magnetic measurements show that FePtAg NPs with a high coercivity of 5.23 kOe can be achieved using the octadecylamine solvent, which is significantly higher than the NPs synthesized using hexadecylamine (2.84 kOe) and trioctylamine (2.81 kOe) solvents. STEM characterization shows that the L10–FePtAg NPs synthesized in octadecylamine, with a diameter of 3.5 ± 0.5 nm and a very uniform size distribution, are smaller in size and more uniform than those synthesized in hexadecylamine and trioctylamine solvents. The octadecylamine, hexadecylamine and trioctylamine high boiling-temperature solvents play significant roles in the morphology, structure and magnetism of FePtAg NPs, and the solvent effects on the nucleation and growth of the L10–FePtAg nanoparticles are proposed. This work sheds new light on designing highly ordered L10–FePt nanoparticles for ultrahigh density magnetic recording and for synthesising high performance oxygen reduction reaction catalysts.

Five new phenolic compounds, gramniphenols C-G (1-5), and eight known compounds (6-13) were isolated from the whole plant of Arundina gramnifolia. Compounds 1, 4, and 5 showed anti-tobacco mosaic virus activity, with IC(50) values of 20.8, 40.8, and 57.7 μM, respectively. Compounds 1-10 were also tested for their anti-HIV-1 activity; compounds 2, 3, and 6 displayed anti-HIV-1 activity with therapeutic index values above 100:1.

Abscisic acid (ABA) is a classical phytohormone which plays an important role in plant stress resistance. Moreover, ABA is also found to regulate the activation of innate immune cells and glucose homeostasis in mammals. Therefore, this ‘stress hormone’ is of great importance to theoretical research and agricultural and medical applications. Botrytis cinerea is a well-known phytopathogenic ascomycete that synthesizes ABA via a pathway substantially different from higher plants. Identification of the functional genes involved in ABA biosynthesis in B. cinerea would be of special interest. We developed an ABA-overproducing mutant strain, B. cinerea TBC-A, previously and obtained a 41.5-Mb genome sequence of B. cinerea TBC-A. In this study, the transcriptomes of B. cinerea TBC-A and its ancestral strain TBC-6 were sequenced under identical fermentation conditions. A stringent comparative transcriptome analysis was performed to identify differentially expressed genes participating in the metabolic pathways related to ABA biosynthesis in B. cinerea. This study provides the first global view of the transcriptional changes underlying the very different ABA productivity of the B. cinerea strains and will expand our knowledge of the molecular basis for ABA biosynthesis in B. cinerea.

RNA tetrahedral nanoparticles with two different sizes are successfully assembled by a one-pot bottom-up approach with high efficiency and thermal stability. The reported design principles can be extended to construct higher-order polyhedral RNA architectures for various applications such as targeted cancer imaging and therapy.

According to the complex nonlinear relationship between gas emission and its effect factors, and the shortcomings that basic colony algorithm is slow, prone to early maturity and stagnation during the search, we introduced a hybrid optimization strategy into a max-min ant colony algorithm, then use this improved ant colony algorithm to estimate the scope of RBF network parameters. According to the amount of pheromone of discrete points, the authors obtained from the interval of network parameters, ants optimize network parameters. Finally, local spatial expansion is introduced to get further optimization of the network. Therefore, we obtain a better time efficiency and solution efficiency optimization model called hybrid improved max-min ant system (HI-MMAS). Simulation experiments, using these theory to predict the gas emission from the working face, show that the proposed method have high prediction feasibility and it is an effective method to predict gas emission.

A series of new chiral side-chain liquid crystalline polysiloxanes(P0-P7) bearing cyano groups were synthesized with a cholesteric LC monomer and a nematic monomer with cyano units(electron withdrawing). The chemical structures and LC properties of the polymers were measured by spectrum and thermal analysis techniques. The obtained polymers were soluble in many solvents and the specific rotations showed negative values and increased with the increasing content of chiral reagent. P1 showed grained texture of smectic A, P2~P7 showed lined texture of chiral smectic C(SC*). XRD curves of P1~P7 samples showed sharp and strong peaks at low angle, but no sharp peak was seen for P0 sample. The temperatures at which 5% weight loss occurred(Td) were greater than 300oC for all the polymers, which reveals all the polymers were stable. And the Tg values of polymers were all low. The Tg and Ti values are all increased with the increasing chiral contents. All the polymers displayed wide range of liquid crystal phase on heating and cooling cycle.

A series of main-chain liquid-crystalline polymers containing maltose were synthesized by interfacial condensation reactions of sebacyl chloride, 4,4’-Dihydroxy-2,2’-dimethyl Benzalazine(DDBA), and 1-O-[4´-O-(α-D-glucopyranosyl)-ß-D-glucopyranosyl]-methanol(MM). The effects of MM on characteristics of liquid-crystalline properties were studied. P1- P3 exhibited nematic mesophase on heating and cooling cycle, while P4- P7 exhibited both chiral smectic A and nematic mesophases on heating cycle and nematic mesophase on cooling cycle. P8 did not exhibit any mesophase on heating and cooling cycle. The glass-transition temperature decreased with increasing concentration of MM.

One-stage pre-cooled mixture refrigerant cycle can be applied in small-scale LNG plant and be special suitable for skit mounted LNG plant. It has different character with the C3MR cycle used in large-scale LNG plant. The optimization of the mixture refrigerant is carried out using HYSYS software. The effect of the main process parameters on the performance of the cycle is calculated and discussed. The result shows that appropriate ranges of the process parameters exist. Higher and lower values of the parameters will increase the energy consumption significantly. The results also indicate that the optimization of the one-stage pre-cooled mixture refrigerant cycle can obtain rather high energy efficiency that is competitive with that of the SMR which is widely employed in small-scale LNG plant.

The synthesis of a new cholesteric monomer (MLC) containing menthyl groups and a series of cholesteric elastomers (LCE1−LCE4) is described. Their chemical structures and purity were characterized by FTIR, 1H-NMR, and elemental analyses. The phase behavior and thermal stability were investigated by differential scanning calorimetry, polarizing optical microscopy, X-ray diffraction, and thermogravimetric analysis. By inserting a flexible spacer between the mesogenic core and the terminal menthyl groups, mesomorphism of MLC was realized. LCE1−LCE4 with low content of crosslinking unit exhibited cholesteric phase because of the introduction of the nematic crosslinking unit. This indicates that low levels of chemical crosslinking do not significantly affect the phase behavior and mesomorphism of the elastomers, and reversible mesophase transitions can be observed. In addition, with increasing the content of crosslinking unit, the corresponding Tg decreased for LCE1−LCE4, whereas their Ti did not remarkable change. © 2012 Wiley Periodicals, Inc. J Appl Polym Sci, 2012

The running condition of a MRC plant is affected by the charge or leakage of the refrigerant. It is significant for the design and operation of the plant. A new model which is established based on the process simulation, mass conservation and characteristics of the system was employed to study the dynamic performance in these cases. The results show that the light composition mainly affects the pressure and the heavy composition affects the liquid level of vessel more obviously. This is due to the fact that the light composition mainly stays in the vapor phase and the heavies stay in the liquid phase mostly. The case when leakages occur at different location was also studied. The results can provide useful information for the adjustment of mixture refrigerant and operation of a MRC plant.

A new kind of bowl-shaped liquid crystals were designed and synthesized by esterification. They used a calix[6]arene as central unit and ω- cholesteric alkyl diacid monoester as mesogenic arms. Their chemical structures were confirmed by FTIR and 1HNMR spectra, their mesomorphic properties and phase behavior were investigated by differential scanning calorimetry (DSC), polarizing optical microscopy (POM) and X-Ray Diffraction (XRD) measurements. The results showed that the kind of calix[6]arene bowl-shaped liquid crystals N1-N3 exhibited broad range liquid crystalline phases at moderate temperature, and they displayed cholesteric phase focal-conic texture in liquid crystal states. With increasing the flexibility of monomers M1-M3, the melting point and the clearing temperature decreased in the macromolecules.

RNA nanoparticles have applications in the treatment of cancers and viral infection; however, the instability of RNA nanoparticles has hindered their development for therapeutic applications. The lack of covalent linkage or crosslinking in nanoparticles causes dissociation in vivo. Here we show that the packaging RNA of bacteriophage phi29 DNA packaging motor can be assembled from 3-6 pieces of RNA oligomers without the use of metal salts. Each RNA oligomer contains a functional module that can be a receptor-binding ligand, aptamer, short interfering RNA or ribozyme. When mixed together, they self-assemble into thermodynamically stable tri-star nanoparticles with a three-way junction core. These nanoparticles are resistant to 8 M urea denaturation, are stable in serum and remain intact at extremely low concentrations. The modules remain functional in vitro and in vivo, suggesting that the three-way junction core can be used as a platform for building a variety of multifunctional nanoparticles. We studied 25 different three-way junction motifs in biological RNA and found only one other motif that shares characteristics similar to the three-way junction of phi29 pRNA.

In the present work, a ester of glucose (a) with 4-(4-(ethoxybenzoyloxy) phenoxy) -6-oxohexanoic acid side–arms has been synthesized. Compared withbandc, reported previously,ahad the same side-arms but different chiral core. The phase behaviors of three star-shaped liquid crystals (a, b, c) have been compared and studied, and the effect of the different structure of chiral core on the phase behaviors of the star-shaped liquid crystals has been discussed. a, b andcdisplayed cholesteric phase, unlike the nematic side arm.a, b andcwere all laevo-rotationary, unlike their parent cores. The melting temperature of a, b andcdecreased with the increase of the sugar ring of the chiral core. The mesomorphic region ofaon cooling cycle was wider than that on heating cycle. Forbandc, the case was similar.

Recent advances in RNA nanotechnology have led to the emergence of a new field and brought vitality to the area of therapeutics [P. Guo, The emerging field of RNA nanotechnology, Nat. Nanotechnol., 2010]. Due to the complementary nature of the four nucleotides and its special catalytic activity, RNA can be manipulated with simplicity characteristic of DNA, while possessing versatile structure and diverse function similar to proteins. Loops and tertiary architecture serve as mounting dovetails or wedges to eliminate external linking dowels. Unique features in transcription, termination, self-assembly, self-processing, and acid-resistance enable in vivo production of nanoparticles harboring aptamer, siRNA, ribozyme, riboswitch, or other regulators for therapy, detection, regulation, and intracellular computation. The unique property of noncanonical base-pairing and stacking enables RNA to fold into well-defined structures for constructing nanoparticles with special functionalities. Bacteriophage phi29 DNA packaging motor is geared by a ring consisting of six packaging RNA (pRNA) molecules. pRNA is able to form a multimeric complex via the interaction of two reengineered interlocking loops. This unique feature makes it an ideal polyvalent vehicle for nanomachine fabrication, pathogen detection, and delivery of siRNA or other therapeutics. This review describes methods in using pRNA as a building block for the construction of RNA dimers, trimers, and hexamers as nanoparticles in medical applications. Methods for industrial-scale production of large and stable RNA nanoparticles will be introduced. The unique favorable PK (pharmacokinetics) profile with a half life (T(1/2)) of 5-10h comparing to 0.25 of conventional 2'-F siRNA, and advantageous in vivo features such as non-toxicity, non-induction of interferons or non-stimulating of cytokine response in animals will also be reviewed.

Deltonin, a steroidal saponin, isolated from Dioscorea zingiberensis Wright (DZW), has shown high-cytotoxic activity in cancer cells. However, its mechanisms and in vivo anti-cancer effects remain unknown. In the present study, we evaluated the effects and explored the anti-tumor mechanisms of deltonin on a panel of colon cancer cell lines and in a mouse model of murine colon cancer C26. Deltonin had more cytotoxic effect on C26 cells than 5-fluorouracil had, promoting dramatic G2-M phase arrest and apoptosis in C26 cells in a concentration-dependent manner; oral administration of deltonin significantly inhibited the tumor growth and prolonged survival of the tumor bearing mice. The deltonin treatment caused a noticeable apoptosis in tumor tissue, which associated with increased levels of Bax, activated caspase-3, caspase-9, and cleaved poly (ADPribose) polymerase, decreased pro-caspase-8, pro-caspase-9, Bcl-2 expression levels and extracellular signal-regulated kinase-1/2 activity; and dose-dependently inhibit angiogenesis. In conclusion, the findings in this study demonstrated that deltonin is an effective natural agent for cancer therapy, which may be mediated, in part, by induction of apoptosis, as well as involve mitogen-activated protein kinase pathways, and inhibition of angiogenesis.

Chiral monomer (M1 ), mesogenic and non-mesogenic crosslinking agents (C1 and C2 ), and the corresponding liquid crystalline elastomers (P1 and P2 series), have been synthesised. Their chemical structures have been characterised by Fourier transform infrared or 1H nuclear magnetic resonance and their phase behaviour investigated by differential scanning calorimetry, polarising optical miscoscopy, thermo-gravimetric analysis (TGA) and X-ray diffraction. The effect of the crosslinking unit on the phase behaviour of the elastomers has been studied. M1 showed a cholesteric oily streak and focal conic texture. C2 exhibited a nematic enantiotropic thread-like and schlieren texture, and a monotropic fan-shaped texture in the SA phase. Due to the introduction of the mesogenic crosslinking unit, elastomers, P2-1 −P2-5 , exhibited a cholesteric phase, while elastomers, P1-1 −P1-4 , derived from a non-mesogenic crosslinking unit, exhibit a SA phase. As the content of the crosslinking unit increased, the T g of the P...

The increasing interest in RNA nanotechnology and the demonstrated feasibility of using RNA nanoparticles as therapeutics have prompted the need for imaging systems with nanometer-scale resolution for RNA studies. Phi29 dimeric pRNAs can serve as building blocks in assembly into the hexameric ring of the nanomotors, as modules of RNA nanoparciles, and as vehicles for specific delivery of therapeutics to cancers or viral infected cells. The understanding of the 3D structure of this novel RNA dimeric particle is fundamentally and practically important. Although a 3D model of pRNA dimer has been proposed based on biochemical analysis, no distance measurements or X-ray diffraction data have been reported. Here we evaluated the application of our customized single-molecule dual-viewing system for distance measurement within pRNA dimers using single-molecule Fluorescence Resonance Energy Transfer (smFRET). Ten pRNA monomers labeled with single donor or acceptor fluorophores at various locations were constructed and eight dimers were assembled. smFRET signals were detected for six dimers. The tethered arm sizes of the fluorophores were estimated empirically from dual-labeled RNA/DNA standards. The distances between donor and acceptor were calculated and used as distance parameters to assess and refine the previously reported 3D model of the pRNA dimer. Distances between nucleotides in pRNA dimers were found to be different from those of the dimers bound to procapsid, suggesting a conformational change of the pRNA dimer upon binding to the procapsid.

Linear double-stranded DNA (dsDNA) viruses package their genome into a procapsid using an ATP-driven nanomotor. Here we report that bacteriophage phi29 DNA packaging motor exercises a one-way traffic property for dsDNA translocation from N-terminal entrance to C-terminal exit with a valve mechanism in DNA packaging, as demonstrated by voltage ramping, electrode polarity switching, and sedimentation force assessment. Without the use of gating control as found in other biological channels, the observed single direction dsDNA transportation provides a novel system with a natural valve to control dsDNA loading and gene delivery in bioreactors, liposomes, or high throughput DNA sequencing apparatus.

A series of side-chain liquid crystalline polysiloxanes bearing different chiral groups were synthesized with a cholesteric liquid crystalline monomer cholesteryl 2-(4-allyloxy-benzoxy)-succinate and a chiral monomer 2-isopropyl-5-methyl-cyclohexyl 4-allyloxy-benzoate. All the chiral polymers showed cholesteric mesophase with very wide mesophase temperature ranges. Along with phase morphology varied from solid state to isotropic state, the sample exhibited varied spectroscopic images, indicating different phase behaviors between mesophase and isotropic state. Optical properties of the polymers were characterized with circular polarization spectra and optical rotation analysis. The specific rotations of the chiral liquid crystalline polymers were all negative values and increased with increase of menthyl component in the polymer systems. As compared with corresponding monomers, the polymers showed significantly lower specific rotations, suggesting that the existence of polysiloxane main chains affect the molecule polarity. The selective light reflection maximum wavelength of the polymers at the same temperature shifted to short wavelength with decrease of cholesteryl component, indicating that the helical pitch becomes shorter due to induction of menthyl groups with high chiral properties. The temperature dependence of selective light reflection maximum wavelength was shifted to a longer wavelength region as the temperature decreased. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009

There is evidence that adenosine acting at A(2A) receptors (A(2A)R) can influence striatal plasticity and cognitive functions. We examined spatial working memory in wild-type (WT) and A(2A) receptor knock-out (KO) mice using two assessments: the eight arm radial maze and a repeated trial Morris water maze (MWM) paradigm. Compared to WT littermates, A(2A)R KO mice displayed enhanced working memory as evidenced by a decrease in escape latency in trial 2 compared to trial 1 in the repeated trial MWM, and by a reduction in working memory errors in the radial arm maze. Both MWM and radial maze results indicated that this enhancement of working memory in A(2A)R KO mice was selective for this specific short-term memory. The decrease in escape latency in MWM was detected with an inter-trial interval of 15 s but not with intervals of 10 or 60 min. In the radial maze, spatial reference memory and memory retention after prolonged training (15 days but not 6 days) were not affected by the A(2A)R KO. These results demonstrate preferential improvement in spatial working memory by genetic inactivation of the A(2A)R and support a modulatory role of the A(2A)R in spatial working memory in mice.

A customized laser combiner was designed and constructed for dual channel single molecule imaging. The feasibility of a combiner-incorporated imaging system was demonstrated in studies of single molecule FRET. Distance rulers made of dual-labeled dsDNA were used to evaluate the system by determining the distance between one FRET pair. The results showed that the system is sensitive enough to distinguish between distances differing by two base pair and the distances calculated from FRET efficiencies are close to those documented in the literature. The single molecule FRET with the dual-color imaging system was also applied to reconstructed phi29 motor pRNA monomers. Finally, techniques for dual laser alignment and tuning of laser power for dual-color excitation are discussed.

Side-chain liquid crystalline ionomer (SLCI) containing sulfonic acid groups with a polymethylhydrosiloxane main-chain was used in the blends of polypropylene (PP) and polybutylene terephthalate (PBT) as a compatibilizer. The crystalline behavior, morphological, and mechanical properties of the blends were investigated in detail by differential scanning calorimetry (DSC), polarizing optical microscope (POM), Fourier transforms infrared spectroscopy (FTIR), and scanning electron microscopy (SEM). Revealed by the shift of Tm in DSC thermogram and the shift of the absorbed peak in FTIR spectra, specific interaction led to stronger interfacial adhesion between these phases, which resulted in much finer dispersion of the minor PBT phase in PP matrix. The SLCI containing sulfonate acid groups acted as physical crosslinking agent along the interface, which compatibilized PP/PBT blends. The mechanical property of the blends including 4 wt % SLCI contents was better than that of other SLCI contents in the blends. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009

Two-component systems (TCSs), typically consisting of a histidine kinase (HK) and a cognate response regulator (RR), are the most common signaling systems in bacteria. Besides paired genes encoding TCSs, there also exists unpaired HKs and orphan RRs. In Streptomyces coelicolor, 13 orphan RRs have been annotated. Because of lack of cognate HKs, little is known as yet about the regulation of orphan RRs. Bioinformatic analysis revealed that several orphan RRs had high amino acid sequence identities with RRs from typical TCSs in S. coelicolor. Among them, the orphan RR SCO3818 and RR SCO0204, which paired with HK SCO0203, showed the highest identity (65%), suggesting that the two RRs might both be under the regulation of SCO0203. Following studies showed that SCO0203 could phosphorylate not only SCO0204 but also SCO3818. Deletion of either sco0203 or sco3818 led to enhanced production of blue-pigmented antibiotic actinorhodin, which indicated a functional correlation between SCO0203 and SCO3818. These results suggested that SCO3818 might be regulated by SCO0203. This is the first report describing the regulation of an orphan RR by an HK. Moreover, this is also the first identification of cross-talk between different TCS components in S. coelicolor.

A series of new chiral side-chain liquid-crystalline polymers were prepared containing smectic-nematic monomer and nonmesogenic chiral monomer. All polymers were synthesized by graft polymerization using polymethylhydrosiloxane as backbone. The mesomorphic properties were investigated by differential scanning calorimetry, polarizing optical microscopy, thermogravimetric analyses, and X-ray diffraction measurements. The chemical structures of the monomers and polymers obtained were confirmed by Fourier transform infrared, proton nuclear magnetic resonance spectra (1H NMR). M1 showed smectic (SB, SC) and nematic phase on the heating and the cooling cycle. Polymers P0–P2 were in chiral smectic A phase, while P3–P5 were in cholesteric phase, P6 has bad LC properties, and P7 has no LC properties. Experimental results demonstrated that nonmesogenic chiral moiety and LC mesogenic with long carbochain offered the possibility of application because of its lower glass transition temperature, and the glass transition temperatures and isotropization temperatures and the ranges of the mesophase temperature reduced with increasing the contents of chiral agent. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2008

To seek more information on function of two-component regulatory systems (TCSs) in Streptomyces coelicolor, a dozen TCS-knockout mutants were generated, and phenotype changes were determined. One TCS (SCO5403/5404)-deleted mutant with phenotype change was obtained. Here, we report the characterization of this novel TCS, designated as RapA1/A2 (regulation of both actinorhodin and a type I polyketide), using genetic and proteomic approaches. Although growth and morphological analyses showed no difference between the knockout mutant and wild-type strain M145, a visible decrease of the production of actinorhodin (Act) was observed in rapA1/A2 mutant. The decrease can be restored by introducing rapA1/A2 genes on an integrative vector. A 2D-gel based proteomic analysis showed that knockout of rapA1/A2 resulted in reduced expression of a putative 3-oxoacyl-[acyl-carrier protein] reductase that is part of a biosynthetic cluster for a cryptic type I polyketide. Further reverse-transcriptase-polymerase chain reaction (RT-PCR) analyses confirmed that expression levels of several biosynthetic genes and the respective pathway-specific regulatory genes actII-ORF4 and kasO for these two clusters were all down-regulated in the rapA1/A2 mutant, compared to M145. Taken together, the results demonstrated that RapA1/A2 may serve as a positive regulator for biosynthesis of both Act and the uncharacterized polyketide in S. coelicolor, and the effects exerted by RapA1/A2 were dependent on the pathway-specific regulatory genes.

Limited by the spatial resolution of optical microscopy, direct detection or counting of single components in biological complexes or nanoparticles is challenging, especially for RNA, which is conformationally versatile and structurally flexible. We report here the assembly of a customized single-molecule dual-viewing total internal reflection fluorescence imaging system for direct counting of RNA building blocks. The RNA molecules were labeled with a single fluorophore by in vitro transcription in the presence of a fluorescent AMP. Precise calculation of identical or mixed pRNA building blocks of one, two, three, or six copies within the bacteriophage phi29 DNA packaging motor or other complexes was demonstrated by applying a photobleaching assay and evaluated by binomial distribution. The dual-viewing system for excitation and recording at different wavelengths simultaneously will enable the differentiation of different complexes with different labels or relative motion of each labeled component in motion machines.

Postoperative pain is the main obstacle for safely rapid recovery of patients undergoing laparoscopic cholecystectomy (LC). In this study, we systemically evaluated the analgesic efficacy of intraperitoneal and incisional ropivacaine injected at the end of the LC. A total of 160 patients, scheduled for elective LC, were allocated into four groups. Group Sham received intraperitoneal and incisional normal saline (NS). Group IC received incisional ropivacaine and intraperitoneal NS. Group IP received incisional NS and intraperitoneal ropivacaine. Group ICP received intraperitoneal and incisional ropivacaine. At the end of the surgery, ropivacaine was injected into the surgical bed through the right subcostal port and infiltrated at the four ports. Dynamic pain by a visual analogue scale (VAS) and cumulative morphine consumption at 2 h, 6 h, 24 h, and 48 h postoperatively, as well as incidence of side-effects over 48 h after LC was recorded. Compared with those in group Sham, the time of post-anesthesia care unit (PACU) stay, dynamic VAS score (VAS-D) 2 h and 6 h postoperatively, cumulative morphine consumption 6 h and 24 h postoperatively, and incidence of nausea and vomiting 48 h after LC in group IC and ICP were less (P

Aims: To find methods for potent drug development by targeting to biocomplex with high copy number. Methods: Phi29 DNA packaging motor components with different stoichiometries were used as model to assay virion assembly with Yang Hui's Triangle [Formula: see text], where Z = stoichiometry, M = drugged subunits per biocomplex, p and q are the fraction of drugged and undrugged subunits in the population. Results: Inhibition efficiency follows a power function. When number of drugged subunits to block the function of the complex K = 1, the uninhibited biocomplex equals qz, demonstrating the multiplicative effect of stoichiometry on inhibition with stoichiometry 1000 > 6 > 1. Complete inhibition of virus replication was found when Z = 6. Conclusion: Drug inhibition potency depends on the stoichiometry of the targeted components of the biocomplex or nanomachine. The inhibition effect follows a power function of the stoichiometry of the target biocomplex.

Gastric cancer is the second leading cause of cancer-related death worldwide. RNA nanotechnology has recently emerged as an important field due to recent finding of its high thermodynamic stability, favorable and distinctive in vivo attributes. Here we reported the use of the thermostable three-way junction (3WJ) of bacteriophage phi29 motor pRNA to escort folic acid, a fluorescent image marker and BRCAA1 siRNA for targeting, imaging, delivery, gene silencing and regression of gastric cancer in animal models. In vitro assay revealed that the RNA nanoparticles specifically bind to gastric cancer cells and knock-down the BRCAA1 gene. Apoptosis of gastric cancer cells was observed. Animal trials confirmed that these RNA nanoparticles could be used to image gastric cancer in vivo, while showing little accumulation in crucial organs and tissues. The volume of gastric tumors noticeably decreased during the course of treatment. No damage to important organs by RNA nanoparticles was detectible. All the results indicated that this novel RNA nanotechnology can overcome conventional cancer therapeutic limitations and opens new opportunities for specific delivery of therapeutics to stomach cancer without damaging normal cells and tissues, reduce the toxicity and side effect, improve the therapeutic effect and exhibit great potential in clinical tumor therapy.

RNA nanotechnology is an emerging field at the interface of biochemistry and nanomaterials that shows immense promise for applications in nanomedicines, therapeutics and nanotechnology. Noncoding RNAs, such as siRNA, miRNA, ribozymes, and riboswitches, play important roles in the regulation of cellular processes. They carry out highly specific functions on a compact and efficient footprint. The properties of specificity and small size make them excellent modules in the construction of multifaceted RNA nanoparticles for targeted delivery and therapy. Biological activity of RNA molecules, however, relies on their proper folding. Therefore their thermodynamic and biochemical stability in the cellular environment is critical. Consequently, it is essential to assess global fold and intracellular lifetime of multifaceted RNA nanoparticles to optimize their therapeutic effectiveness. Here, we describe a method to express and assemble stable RNA nanoparticles in cells, and to assess the folding and turnover rate of RNA nanoparticles in vitro as well as in vivo in real time using a thermostable core motif derived from pRNA of bacteriophage Phi29 DNA packaging motor and fluorogenic RNA modules.

In order to efficiently introduce DNA into B. subtilis ZK, which produces iturin A at a high level, we optimized seven electroporation conditions and explored an efficient electroporation method. Using the optimal conditions, the electroporation efficiency was improved to 1.03 × 107 transformants/μg of DNA, an approximately 10,000-fold increase in electroporation efficiency. This efficiency is the highest electroporation efficiency for B. subtilis and enables the construction of a directed evolution library or the knockout of a gene in B. subtilis ZK for molecular genetics studies. In the optimization process, the combined effects of three types of wall-weakening agents were evaluated using a response surface methodology (RSM) design, which led to a two orders of magnitude increase in electroporation efficiency. To the best of our limited knowledge, this study provides the first demonstration of using an RSM design for optimization of the electroporation conditions for B. subtilis. To validate the electroporation efficiency, a case study was performed and a gene (rapC) was inactivated in B. subtilis ZK using a suicide plasmid pMUTIN4. Moreover, we found that the rapC mutants exhibited a marked decrease in iturin A production, suggesting that the rapC gene was closely related to the iturin A production.

A series of siloxane‐based liquid crystalline elastomers containing biphenyl benzoate mesogenic units and ionic Brilliant Yellow moieties was synthesized. The chemical structures and liquid crystalline properties of the samples were characterized by FTIR, 1H NMR, DSC, POM and XRD. The effective crosslink density of the ionic elastomers was determined by swelling experiments in organic/buffer mixtures. All the polymers displayed a smectic mesophase. It was shown that the glass transition and melting point temperatures of the polymers increased slightly with increasing content of ionic and mesogenic groups in the polymers, while the liquid crystal mesophase region decrease slightly.

Four bisacrylate mesogenic monomers and the corresponding liquid-crystalline thermosets were synthesized. The chemical structures of the intermediate compounds and monomers obtained were confirmed by elemental analyses, Fourier transform infrared, and 1H NMR and 13C NMR spectra. The mesomorphic properties and thermal stability were investigated with differential scanning calorimetry, thermogravimetric analysis, polarized optical microscopy, and X-ray diffraction measurements. The influence of the curing temperatures and time on the phase behavior and thermal stability of the thermosets was discussed. All the monomers and thermosets exhibited a nematic schlieren texture. However, the monomers only showed the melting transition, and the thermosets displayed the glass transition. The experimental results demonstrated that the monomer structures strongly affected the phase behavior and the curing reaction rate, and the glass-transition temperatures and thermal stability of the thermosets increased with the curing temperature and time. © 2005 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 43: 4478–4485, 2005

A new class of star-shaped glassy nematic liquid crystals based on pentaerythritol as a flexible core and ω-[4-(p-alkoxybenzoloxy)phenoxycarbonyl]valeric acid as the side-chain mesogenic arms has been prepared. Their structures and mesomorphism have been investigated in detail.

A mesogenic crosslinking agent M-1 was synthesized to minimize the perturbations of non-mesogenic crosslinking agents in liquid crystalline elastomers. The synthesis of new side chain liquid crystalline elastomers containing the rigid mesogenic crosslinking agent M-1 and nematic monomer M-2 by a one-step hydrosilylation reaction is described. The chemical structures of the monomers and network polymers obtained were confirmed by FTIR and 1H NMR spectroscopy. The mesomorphic properties and phase behaviour were investigated by differential scanning calorimetry, polarizing optical microscopy, and X-ray diffraction. The influence of the crosslinking units on phase behaviour is discussed. Liquid crystalline elastomers containing less than 15 mol % of the crosslinking units showed elasticity, reversible phase transitions and a threaded texture. The experimental results demonstrated that the glass transition temperature of polymers P-1–7 increased with increasing concentration of crosslinking agent M-1; but the ...

The synthesis of six liquid crystalline monomers M1–M6 and three series of side chain cholesteric liquid crystalline polymers P1–P3 is described. The chemical structures of the monomers were characterized by FTIR and 1H NMR spectroscopy. The structure–property relationships of M1–M6 and P1–P3 are discussed. Their phase behaviour and optical properties were investigated by differential scanning calorimetry and polarizing optical microscopy. The monomers M1–M3 exhibited cholesteric phases; M4–M6 showed nematic or smectic phases. The polymer series P1–P3 showed cholesteric phases. Experimental results demonstrated that the selective reflection of the cholesteric monomers and homopolymers shifted to the shorter wavelength region (blue shift) with increasing length of the flexible spacer. The selective reflection of the copolymers shifted to the longer wavelength region (red shift) with increasing content of nematic units.

The synthesis of hemiphasmidic monomers 4-[(3,4,5-triethoxy)benzoyloxy]-4′-[(p-allyloxy)benzoyloxy]biphenyl (M1), 4-[(3,5-diethoxy)benzoyloxy]-4′-[(p-allyloxy)-benzoyloxy]biphenyl (M2), and of the corresponding side-chain liquid-crystalline polysiloxanes (P1, P2) was carried out. For comparison, rodlike monomer 4-[(p-ethoxy)-benzoyloxy]-4′-[(p-allyloxy)benzoyloxy]biphenyl (M3) and its polysiloxanes (P3) were also prepared. The chemical structures of the monomers and polymers obtained were confirmed by FTIR and 1H-NMR spectra. Their mesomorphic properties and phase behavior were investigated by differential scanning calorimetry, polarizing optical microscopy, and X-ray diffraction measurements. The relationship between structures and properties was discussed. It was observed that M1 and M3 were enantiotropic nematic phase, M2 was monotropic mesophase, and their poly(methylsiloxanes) (P1–P3) possessed a broad range enantiotropic nematic phases and high thermal stability. © 2004 Wiley Periodicals, Inc. J Appl Polym Sci 95: 946–952, 2005

In nanotechnology, biomolecular assemblies serve not only as model systems for the construction of nanodevices, but they can also be used directly as templates for the formation of nanostructures. Biological nano-building blocks can either be isolated as complete functional units from living cells or viruses (biological "Top down" approach) or formed by biomolecular assembly from recombinant or synthetic components ("Bottom up" approach). In both cases, rational design of nanostructures requires knowledge of the stoichiometry of the biological structures, which frequently occur as multimers, i.e., the morphological complex is composed of multiple copies of one or more macromolecules. In this paper, a method is described for the stoichiometric quantification of molecules in bio-nanostructures. The method is based on using dilution factors and relative concentrations rather than absolute quantities, which are often difficult to determine, especially in short-lived assembly intermediates. The approach exploits the fact that the larger the stoichiometry of the component is, the more dramatic is the influence of the dilution factor (decrease in concentration) on the reaction. We established and used the method to determine the stoichiometry of components of bacterial virus phi29. The log of dilution factors was plotted against the log of reaction yield. The stoichiometry Z was determined with the equation Z=-1.58+2.4193T-0.001746T(2) [T in (0,1000), or 90 degree angle alpha in (0 degrees, 89.9 degrees )], where T is the slope of the curve (tangent of 90 degree angle alpha, which is the angle between the x-axis and the concentration dependent curve). Z can also be determined from a standard table given in this report. With the bacteriophage phi29 in vitro assembly system, up to 5x10(8) infectious virions per ml can be assembled from 11 purified components, giving our method a sensitivity of nine orders of magnitude. We confirmed the stoichiometries of phi29 components that were determined previously with microscopic approaches. The described method also responded to programmed stoichiometry changes, which were generated by assembling the phi29 DNA packaging motor from modified pRNA (DNA-packaging RNA) molecules forming a trimer of dimers or a dimer of trimers, instead of the wild-type hexamer.

Key Laboratory of Tobacco Chemistry of Yunnan Province, Yunnan Academy of Tobacco Science, Kunming 650106, P.R. ChinaReceived May 26, 2012, Accepted October 2, 2012Key Words : Rosa rugosa, Phenylethanoids, Anti-HIV-1 activity, Cytotoxic abilityRosa rugosa is widely distributed from temperate regionsof eastern Asia including Japan, Korea and China.

Background: Few clinical trials have evaluated the efficacy and safety of Tripterygium wilfordii Hook F (TwHF) compared with acitretin in psoriasis. We aimed to compare the efficacy and safety of TwHF compared with acitretin in the treatment of moderate to severe psoriasis vulgaris. Methods: Adults with Psoriasis Area Severity Index (PASI) score ≥ 10 and psoriasis-affected body surface area ≥ 10% were randomized into a TwHF (20 mg, 3 times a day) or acitretin group (30 mg, once a day). The treatment course lasted for 8 weeks. Patients were assessed at baseline and at 2, 4, and 8 weeks. Laboratory tests were performed at baseline, week 4, and week 8. The data were analyzed using paired samples t-test or analysis of variance (ANOVA). Results: A total of 115 patients was enrolled (58 TwHF; 57 acitretin). The median PASI score improved in the TwHF group by 50.4% and in the acitretin group by 42.7%. There was no significant difference in median PASI improvement between two groups at 2, 4, and 8 weeks. There was also no significant difference in PASI 25, PASI 50, PASI 75, and PASI 90 response between the two groups at 2, 4, and 8 weeks. There was a significant increase in the level of aspartate transaminase and triglycerides in the TwHF group (P = 0.026 and P = 0.011, respectively). In the acitretin group, there was a significant increase in the level of alanine transaminase, cholesterol, and high-density lipoprotein (P = 0.030, P < 0.01, and P < 0.01, respectively). Conclusions: There was no significant difference in treatment efficacy between the TwHF and acitretin groups within 8 weeks, but there were fewer treatment-related adverse events in the TwHF group.