CRISPRimmunity: an interactive web server for CRISPR-associated Important Molecular events and Modulators Used in geNome edIting Tool identifYing.

The CRISPR-Cas system is a highly adaptive and RNA-guided immune system found in bacteria and archaea, which has applications as a genome editing tool and is a valuable system for studying the co-evolutionary dynamics of bacteriophage interactions. Here introduces CRISPRimmunity, a new web server designed for Acr prediction, identification of novel class 2 CRISPR-Cas loci, and dissection of key CRISPR-associated molecular events. CRISPRimmunity is built on a suite of CRISPR-oriented databases providing a comprehensive co-evolutionary perspective of the CRISPR-Cas and anti-CRISPR systems. The platform achieved a high prediction accuracy of 0.997 for Acr prediction when tested on a dataset of 99 experimentally validated Acrs and 676 non-Acrs, outperforming other existing prediction tools. Some of the newly identified class 2 CRISPR-Cas loci using CRISPRimmunity have been experimentally validated for cleavage activity in vitro. CRISPRimmunity offers the catalogues of pre-identified CRISPR systems to browse and query, the collected resources or databases to download, a well-designed graphical interface, a detailed tutorial, multi-faceted information, and exportable results in machine-readable formats, making it easy to use and facilitating future experimental design and further data mining. The platform is available at http://www.microbiome-bigdata.com/CRISPRimmunity. Moreover, the source code for batch analysis are published on Github (https://github.com/HIT-ImmunologyLab/CRISPRimmunity).

Numerical implementation of three-dimensional vectorial complex ray model and application to rainbow scattering of spheroidal drops.

The rainbow patterns of oblate spheroidal drops have been observed in experiments nearly forty years ago [Nature312, 529 (1984)10.1038/312529a0]. However, the prediction for those complex patterns has been a challenge for conventional light scattering models. The vectorial complex ray model (VCRM) allows to account for the direction, the polarization, the phase, the amplitude and the wavefront curvature of waves and provides a powerful tool for the study of the light/electromagnetic wave interaction with a homogeneous object of any shape with smooth surface. In [Opt. Lett.46, 4585 (2021)10.1364/OL.434149], the authors have reported an important breakthrough of VCRM for the three-dimensional scattering (VCRM3D) and the simulated rainbow patterns of oblate drops. The present paper is devoted to the detailed description of the numerical implementation allowing the simulation of the 3D scattering field by a nonspherical particle. Its ability to predict both the fine and coarse intensity structures of the rainbows and the near-backward scattering patterns of spheroids is demonstrated. This work opens perspectives for exploring the 3D scattering characteristics of large objects with any smooth shape and developing relevant optical techniques for particle characterization.

Development of a quasi-coaxis dual-energy flat spectral response X-ray imaging instrument for measuring hotspot electron temperature.

The measurement of hotspot electron temperature is a paramount technique of implosion physics research in inertial confinement fusion. This study proposes a novel quasi-coaxis dual-energy flat spectral response high-resolution X-ray imaging instrument comprising a dual-channel total-reflection Kirkpatrick-Baez microscope and two flat non-periodic multilayer mirrors, which can image at 6.4 ± 0.5 and 9.67 ± 0.5 keV simultaneously. Various theoretical simulations were performed to verify the performance and feasibility of the imaging instrument, which was assembled and characterized in a laboratory. Experimental results show that the imaging instrument could achieve a high spatial resolution of 5 µm in a ± 150 µm field of view (FOV), the root mean square(RMS) deviation values of the measured reflection efficiency are 1.71% and 1.82% for the 6.4 keV and 9.67 keV imaging channels, respectively, in the ± 150 µm FOV.

Airy theory revisited with the method combining vectorial complex ray model and physical optics.

Airy published his theory in the 1830s to remedy the problem of infinite intensity in the rainbow angles of a spherical droplet predicted by geometrical optics. This theory has been studied by mathematicians and physicists since then from different points of view. In what concerns the scattering diagram around the rainbow angles, Airy theory has been improved by researchers in order to predict correctly the intensity of the supernumerary bows. However, it is known that the positions and the intensities of the supernumerary bows predicted by Airy theory differ from those of rigorous Debye theory with increasing order p and the scattering angles from the rainbow angle. In the present Letter, we will show that this discrepancy is caused by the approximations in Airy theory and can be revised by combining the vectorial complex ray model and physical optics (PO). The former permits us to calculate rigorously the amplitudes and phases of all rays and predicts precisely the scattering pattern except for the main bows. The combination with PO predicts very precisely all the supernumerary bows for both perpendicular and parallel polarization. This method can be applied directly to the light scattering of non-spherical particles with smooth surfaces.

Measurement of Time-Dependent Drive Flux on the Capsule for Indirectly Driven Inertial Confinement Fusion Experiments.

A new method for measuring the time-dependent drive flux at the hohlraum center is proposed as a better alternative to conventional wall-based techniques. The drive flux here is obtained by simultaneous measurement of the reemitted flux and shock velocity from a three-layered "cakelike" sample. With these two independent observables, the influence induced by the uncertainty of the material parameters of the sample can be effectively decreased. The influence from the closure of the laser entrance hole, which was the main challenge in conventional wall-based techniques, was avoided through localized reemitted flux measurement, facilitating drive flux measurement throughout the entire time history. These studies pave a new way for probing the time-dependent drive flux, for both cylindrical hohlraums and novel hohlraums with six laser entrance holes.

The crystal structure of Cpf1 in complex with CRISPR RNA.

The CRISPR-Cas systems, as exemplified by CRISPR-Cas9, are RNA-guided adaptive immune systems used by bacteria and archaea to defend against viral infection. The CRISPR-Cpf1 system, a new class 2 CRISPR-Cas system, mediates robust DNA interference in human cells. Although functionally conserved, Cpf1 and Cas9 differ in many aspects including their guide RNAs and substrate specificity. Here we report the 2.38 Å crystal structure of the CRISPR RNA (crRNA)-bound Lachnospiraceae bacterium ND2006 Cpf1 (LbCpf1). LbCpf1 has a triangle-shaped architecture with a large positively charged channel at the centre. Recognized by the oligonucleotide-binding domain of LbCpf1, the crRNA adopts a highly distorted conformation stabilized by extensive intramolecular interactions and the (Mg(H2O)6)(2+) ion. The oligonucleotide-binding domain also harbours a looped-out helical domain that is important for LbCpf1 substrate binding. Binding of crRNA or crRNA lacking the guide sequence induces marked conformational changes but no oligomerization of LbCpf1. Our study reveals the crRNA recognition mechanism and provides insight into crRNA-guided substrate binding of LbCpf1, establishing a framework for engineering LbCpf1 to improve its efficiency and specificity for genome editing.

Generalized rainbow patterns of oblate drops simulated by a ray model in three dimensions.

The scattering patterns near the primary rainbow of oblate drops are simulated by extending the vectorial complex ray model (VCRM) [Opt. Lett.36, 370 (2011)OPLEDP0146-959210.1364/OL.36.000370] to three-dimensional (3D) calculations. With the curvature of a wavefront as an intrinsic property of a ray, this advanced ray model permits, in principle, to predict the amplitudes and phases of all emergent rays with a rigorous algebraic formalism. This Letter reports a breakthrough of VCRM for 3D scattering with a line-by-line triangulation interpolation algorithm allowing to calculate the total complex amplitude of a scattered field. This makes possible to simulate not only the skeleton (geometrical rainbow angles, hyperbolic-umbilic caustics), but also the coarse (Airy bows, lattice) and fine (ripple fringes) structures of the generalized rainbow patterns (GRPs) of oblate drops. The simulated results are found qualitatively and quantitatively in good agreement with experimental scattering patterns for drops of different aspect ratios. The physical interpretation of the GRPs is also given. This work opens up prominent perspectives for simulating and understanding the 3D scattering of large particles of any shape with a smooth surface by VCRM.

A thermophilic chitinase 1602 from the marine bacterium Microbulbifer sp. BN3 and its high-level expression in Pichia pastoris.

Chitinases play an important role in many industrial processes, including the preparation of oligosaccharides with potential applications. In the present study, a 1,713 bp gene of Chi1602, derived from a marine bacterium Microbulbifer sp. BN3, encoding a GH18 family chitinase, was expressed at high levels in Pichia pastoris. Distinct from most of the marine chitinases, the recombinant chitinase 1602 exhibited maximal activity at 60 °C and over a broad pH range between 5.0 and 9.0, and was stable at 50 °C and over the pH range 4.0-9.0. The hydrolytic products derived from colloidal chitins comprised mainly (GlcNAc)2 and GlcNAc, indicating that rChi1602 is a GH18 processive chitinase in conformity with its hypothetical structure. However, rChi1602 showed traces of ß-N-acetylglucosaminidase activity on substrates such as powder chitin, chitosan, and ethylene glycol chitin. The thermophilic rChi1602, which manifests adaptation to a wide pH range and can be expressed at a high level in P. pastoris, is advantageous for applications in industrial processes.

Development of a polar-view Kirkpatrick-Baez X-ray microscope for implosion asymmetry studies.

The development of a polar-view Kirkpatrick-Baez microscope, fielded in the upper polar zone of the Shenguang-III laser fusion facility, is presented. With this microscope, the resolving power of polar-direction X-ray imaging diagnostics is improved, to the 3 ~5 µm scale. The microscope is designed for implosion asymmetry studies, with response energy points at 1.2 keV, 3.5 keV, and 8 keV. A biperiodic multilayer scheme is adopted to accommodate multiple implosion stages. We present the overall optical system design, target aiming scheme, characteristic composite imaging diagnostic experiments and initial results. The inertial-driven quasi-one-dimensional spherical implosions were observed from orthogonal directions with a convergence ratio of ~14.4. Fine features of the stagnating hot spot core are also resolved.

Three new sesquiterpenes from Pterocarpus santalinus.

Three new sesquiterpenes of canusesnol K (1), canusesnol L (2) and 12, 15-dihydroxycurcumene (3), along with five known ones (4-8), were isolated from the heartwood extract of Pterocarpus santalinus. Their structures were established by extensive analyses of 1D and 2D NMR spectroscopy, including 1H NMR, 13C NMR, HSQC, HMBC and NOESY, and HRESI-MS. The absolute configurations of the new compounds were established with Modified Mosher's method. The cytotoxic activities of all these compounds against HepG2 (human liver cancer), MCF-7 (human breast cancer), MDA-MB-231 (human breast cancer), and Hela (human cervical carcinoma) cancer cell lines were evaluated. Compound 1 exhibited moderate cytotoxic activity toward MDA-MB-231 cell lines.

Computation of radiation pressure force exerted on arbitrary shaped homogeneous particles by high-order Bessel vortex beams using MLFMA.

Due to special characteristics of nondiffraction and self reconstruction, the Bessel beams have attracted wide attention in optical trapping and appear to be a dramatic alternative to Gaussian beams. We present in this paper an efficient approach based on the surface integral equations (SIE) to compute the radiation pressure force (RPF) exerted on arbitrary shaped homogeneous particles by high-order Bessel vortex beam (HOBVB). The incident beam is described by vector expressions perfectly satisfy Maxwell's equations. The problem is formulated with the combined tangential formulation (CTF) and solved iteratively with the aid of the multilevel fast multipole algorithm (MLFMA). Then RPF is computed by vector flux of the Maxwell's stress tensor over a spherical surface tightly enclosing the particle and analytical expression for electromagnetic fields of incident beam in near region are used. The numerical predictions are compared with the results of the rigorous method for spherical particle to validate the accuracy of the approach. Some numerical results on relative large particles of complex shape, such as biconcave cell-like particles with different geometry parameters are given, showing powerful capability of our approach. These results are expected to provide useful insights into the RPF exerted on complex shaped particles by HOBVB.

Experimental validation of the vectorial complex ray model on the inter-caustics scattering of oblate droplets.

We report the first experimental validation of the Vectorial Complex Ray Model (VCRM) using the scattering patterns of large oblate droplets trapped in an acoustic field. The two principal radii and refractive index of the droplets are retrieved with a minimization method that involves VCRM predictions and experimental light scattering patterns. The latter are recorded in the droplet equatorial plane between the primary rainbow region and the associated hyperbolic-umbilic diffraction catastrophe. The results demonstrate that the VCRM can predict the fine and coarse stuctures of scattering patterns with good precision, opening up perspectives for the characterization of large non-spherical particles.

Computational study of radiation torque on arbitrary shaped particles with MLFMA.

The surface integral equation (SIE) method is used for the computational study of radiation torque on arbitrarily shaped homogeneous particles. The Multilevel Fast Multipole Algorithm (MLFMA) is employed to reduce memory requirements and improve the capability of SIE. The resultant matrix equations are solved iteratively to obtain equivalent electric and magnetic currents. Then, radiation torque is computed using the vector flux of the pseudotensor over a spherical surface tightly enclosing the particle. We use, therefore, the analytical electromagnetic field expression for incident waves in the near region, instead of the far-field approximation. This avoids the error which may be caused when describing the incident beam. The numerical results of three kinds of non-spherical particles are presented to illustrate the validity and capability of the developed method. It is shown that our method can be applied to predict, in the rigorous sense, the torque from a beam of any shape on a particle of complex configuration with a size parameter as large as 650. The radiation torques on large ellipsoids are exemplified to show the performance of the method and to study the influence that different aspect ratios have on the results. Then, the code is used for the calculation of radiation torque on objects of complex shape including a biconcave cell-like particle and a motor with a non-smooth surface.

Direct measurement of x-ray flux for a pre-specified highly-resolved region in hohlraum.

A space-resolving flux detector (SRFD) is developed to measure the X-ray flux emitted from a specified region in hohlraum with a high resolution up to 0.11mm for the first time. This novel detector has been used successfully to measure the distinct X-ray fluxes emitted from hot laser spot and cooler re-emitting region simultaneously, in the hohlraum experiments on SGIII prototype laser facility. According to our experiments, the ratio of laser spot flux to re-emitted flux shows a strong time-dependent behavior, and the area-weighted flux post-processed from the measured laser spot flux and re-emitting wall flux agrees with that measured from Laser Entrance Hole by using flat-response X-ray detector (F-XRD). The experimental observations is reestablished by our two-dimensional hydrodynamic simulations and is well understood with the power balance relationship.

Highly efficient expression and characterization of a ß-mannanase from Bacillus subtilis in Pichia pastoris.

A ß-mannanase gene (Man5) from Bacillus subtilis BS5 was cloned by PCR and integrated into the genome of Pichia pastoris GS115 via pPIC9 vector. The recombinant Man5 with a molecular mass of 43 kDa was successfully expressed and secreted into the culture medium. After methanol induction in a shake flask for 96 H, the recombinant Man5 protein reached 375 µg/mL in concentration, with an enzyme activity of 892 U/mL. The recombinant Man5 was purified 3.35-fold with 60% yield by using HiTrap DEAE FF and HiTrap Phenyl FF columns. The specific activity of the purified enzyme was 7,978 U/mg. The optimum temperature and pH of the recombinant Man5 were 50 °C and 6.0, respectively. Studies of substrate specificity showed that the optimum substrate for the Man5 was konjac flour, suggesting that it has great potential as an effective additive in the food industry.

Discovery and structural mechanism of DNA endonucleases guided by RAGATH-18-derived RNAs.

CRISPR-Cas systems and IS200/IS605 transposon-associated TnpBs have been utilized for the development of genome editing technologies. Using bioinformatics analysis and biochemical experiments, here we present a new family of RNA-guided DNA endonucleases. Our bioinformatics analysis initially identifies the stable co-occurrence of conserved RAGATH-18-derived RNAs (reRNAs) and their upstream IS607 TnpBs with an average length of 390 amino acids. IS607 TnpBs form programmable DNases through interaction with reRNAs. We discover the robust dsDNA interference activity of IS607 TnpB systems in bacteria and human cells. Further characterization of the Firmicutes bacteria IS607 TnpB system (ISFba1 TnpB) reveals that its dsDNA cleavage activity is remarkably sensitive to single mismatches between the guide and target sequences in human cells. Our findings demonstrate that a length of 20 nt in the guide sequence of reRNA achieves the highest DNA cleavage activity for ISFba1 TnpB. A cryo-EM structure of the ISFba1 TnpB effector protein bound by its cognate RAGATH-18 motif-containing reRNA and a dsDNA target reveals the mechanisms underlying reRNA recognition by ISFba1 TnpB, reRNA-guided dsDNA targeting, and the sensitivity of the ISFba1 TnpB system to base mismatches between the guide and target DNA. Collectively, this study identifies the IS607 TnpB family of compact and specific RNA-guided DNases with great potential for application in gene editing.

Computation of radiation pressure force on arbitrary shaped homogenous particles by multilevel fast multipole algorithm.

A full-wave numerical method based on the surface integral equation for computing radiation pressure force (RPF) exerted by a shaped light beam on arbitrary shaped homogenous particles is presented. The multilevel fast multipole algorithm is employed to reduce memory requirement and to improve its capability. The resultant matrix equation is solved by using an iterative solver to obtain equivalent electric and magnetic currents. Then RPF is computed by vector flux of the Maxwell's stress tensor over a spherical surface tightly enclosing the particle. So the analytical expressions for electromagnetic fields of incident beam in near region are used. Some numerical results are performed to illustrate the validity and capability of the developed method. Good agreements between our method and the Lorenz-Mie theory for spherical and small spheroidal particle are found while our method has powerful capability for computing RPF of any shaped beam on a relatively large particle of complex shape. Tests for ellipsoidal and red blood cell-like particles illuminated by Gaussian beam have shown that the size of the particle can be as large as 50-100 wavelengths, respectively, for the relative refractive of 1.33 and 1.1.

Scattering of a Gaussian beam by an elliptical cylinder using the vectorial complex ray model.

The scattered waves of a shaped beam by an infinite cylinder in the far field are, stricto sensu, neither cylindrical nor spherical, so the asymptotic form of special functions involved in the theories based on the rigorous solution of Maxwell equations cannot be used to evaluate scattered intensities, even in the most simple case of Gaussian beam scattering by an infinite circular cylinder. Thus, although theories exist for the scattering of a shaped beam by infinite cylinders with circular and elliptical sections, the numerical calculations are limited to the near field. The vectorial complex ray model (VCRM) developed by Ren et al. describes waves by rays with a new property: the curvature of the wavefront. It is suitable to deal with the scattering of an arbitrarily shaped beam by a particle with a smooth surface of any form. In this paper, we apply this method to the scattering of an infinite elliptical cylinder illuminated by a Gaussian beam at normal incidence with an arbitrary position and orientation relative to the symmetric axis of the elliptical section of the cylinder. The method for calculating the curvature of an arbitrary surface is given and applied in the determination of the two curvature radii of the Gaussian beam wavefront at any point. Scattered intensities for different parameters of the beam and the particle as well as observation distance are presented to reveal the scattering properties and new phenomena observed in the beam scattering by an infinite elliptical cylinder.

Double-bundle reconstruction results in superior clinical outcome than single-bundle reconstruction.

PURPOSE: To compare the short- and long-term clinical outcomes of the double-bundle (DB) anterior cruciate ligament (ACL) reconstruction with those of single-bundle (SB) ACL reconstruction. METHODS: An electronic search of the database PubMed (1966-September 2011), EMBASE (1984-September 2011), and Cochrane Controlled Trials Register (CENTRAL; 3rd Quarter, 2011) was undertaken to identify relevant studies. Main clinical outcomes were knee stability measurements including KT-1000 arthrometer measurement, Pivot shift test, and Lachman test, and clinical outcome measurements including International Knee Documentation Committee (IKDC), Lysholm knee score, Tegner activity score, and complications. RESULTS: Eighteen studies were finally included in this meta-analysis, which were all classified as high risk of bias according to the Collaboration's recommended tool. It is seen that compared to SB ACL reconstruction, DB ACL reconstruction results in a KT-1000 arthrometer outcome 0.63 and 1.00 mm closer to the normal knee in a short- and long-term follow-up, respectively. Our results also reveal that DB-treated patients have a significantly higher negative rate of the pivot shift test (p < 0.00001 and = 0.006 in a short- and long-term follow-up, respectively) and Lachman test (n.s. and p < 0.0001 in a short- and long-term follow-up, respectively) compared to SB-treated patients. As for the clinical outcome measurements, a significant difference is found between SB versus DB ACL reconstruction regarding the IKDC (p = 0.006 and < 0.0001 in a short- and long-term follow-up, respectively) and complications (p = 0.03), while there is no significant difference between the two groups regarding Lysholm knee score (n.s.) and Tegner activity score (n.s.). CONCLUSION: Overall, double-bundle ACL reconstruction yields better clinical outcomes when compared to single-bundle ACL reconstruction. LEVELS OF EVIDENCE: II.

Interleukin-18 promoter polymorphism and asthma risk: a meta-analysis.

Some studies have shown that IL-18 was associated with aetiology and progression of asthma. However, the association between single-nucleotide polymorphisms -607C/A (rs1946518) and -137G/C (rs187238) located in the IL-18 gene promoter and asthma risk was still controversial and ambiguous. To derive a more precise effect on the association between these polymorphisms and asthma risk, we performed a meta-analysis based on the currently available evidence of the literature. A total of 5 studies with 1411 cases and 1525 controls for -607C/A polymorphism and 5 studies with 1883 cases and 6645 controls for -137G/C polymorphism were identified to perform a meta-analysis, up to October 2010. Summary ORs and corresponding 95% CIs for IL-18 polymorphisms and asthma were estimated using fixed- and random-effects models when appropriate. Heterogeneity and publication bias were evaluated. We found that individuals carrying AC/CC genotype of -607C/A polymorphism were associated with an increased asthma risk in recessive model (OR = 1.278; 95% CI, 1.073-1.522). However, no significant association was observed between -137G/C polymorphism and asthma risk under different contrast models. There was no evidence of publication bias. The present meta-analysis suggested that IL-18 -607C/A polymorphism in promoter region was associated with asthma risk.