Unmodificated stepless regulation of CRISPR/Cas12a multi-performance.

As CRISPR technology is promoted to more fine-divided molecular biology applications, its inherent performance finds it increasingly difficult to cope with diverse needs in these different fields, and how to more accurately control the performance has become a key issue to develop CRISPR technology to a new stage. Herein, we propose a CRISPR/Cas12a regulation strategy based on the powerful programmability of nucleic acid nanotechnology. Unlike previous difficult and rigid regulation of core components Cas nuclease and crRNA, only a simple switch of different external RNA accessories is required to change the reaction kinetics or thermodynamics, thereby finely and almost steplessly regulating multi-performance of CRISPR/Cas12a including activity, speed, specificity, compatibility, programmability and sensitivity. In particular, the significantly improved specificity is expected to mark advance the accuracy of molecular detection and the safety of gene editing. In addition, this strategy was applied to regulate the delayed activation of Cas12a, overcoming the compatibility problem of the one-pot assay without any physical separation or external stimulation, and demonstrating great potential for fine-grained control of CRISPR. This simple but powerful CRISPR regulation strategy without any component modification has pioneering flexibility and versatility, and will unlock the potential for deeper applications of CRISPR technology in many finely divided fields.

Analysis on the minimum structure of harmonic spectra from MgO crystals.

Recent theoretical and experimental findings have demonstrated the minimum characteristic in the harmonic spectrum of bulk MgO crystals subjected to intense laser pulses. However, the dominant mechanism behind this minimum structure is still under debate. This study simulates the harmonic spectrum from a MgO crystal in a linearly polarized laser pulse by solving multi-band semiconductor Bloch equations. The results show that the minimum feature at 20 eV in the MgO harmonic spectra from 1700 and 800 nm laser pulses is due to band dispersion and interference between interband harmonics. Notably, the disappearance of the minimum structure at 14 eV in the harmonic spectrum from the 800 nm laser is attributed to the intensity suppression of higher energy harmonics, caused by decreased electron population at the boundary of the first Brillouin zone in the multi-band case. These findings offer insights into the spectral structure of solid-state harmonics, contributing to the all-optical reconstruction of the crystal band based on its harmonic spectrum.

A PAM-free CRISPR/Cas12a ultra-specific activation mode based on toehold-mediated strand displacement and branch migration.

CRISPR (clustered regularly interspaced short palindromic repeats) technology has achieved great breakthroughs in terms of convenience and sensitivity; it is becoming the most promising molecular tool. However, only two CRISPR activation modes (single and double stranded) are available, and they have specificity and universality bottlenecks that limit the application of CRISPR technology in high-precision molecular recognition. Herein, we proposed a novel CRISPR/Cas12a unrestricted activation mode to greatly improve its performance. The new mode totally eliminates the need for a protospacer adjacent motif and accurately activates Cas12a through toehold-mediated strand displacement and branch migration, which is highly universal and ultra-specific. With this mode, we discriminated all mismatch types and detected the EGFR T790M and L858R mutations in very low abundance. Taken together, our activation mode is deeply incorporated with DNA nanotechnology and extensively broadens the application boundaries of CRISPR technology in biomedical and molecular reaction networks.

Effect of pulse duration on the above-threshold ionization of a hydrogen atom irradiated by a 400 nm intense laser.

The photoelectron emission spectra generated by the interaction between ultrashort intense laser pulses and atoms can reveal the ultrafast dynamics of electrons. By using the numerical solution of the time-dependent Schrödinger equation in momentum space, the photoelectron emission spectra of atoms irradiated by 400 nm intense lasers with different durations of the pulse has been investigated. In the photoelectron emission spectrum, in addition to the above-threshold ionization peaks due to ionization interference in multiple cycles and the sideband peaks mainly due to the interference of ionized electrons at different moments along the rising edge of the laser pulse envelope, additional peaks of photoelectron emission whose intensity appears to oscillate with the increasing duration of the laser pulse can also be observed. Based on strong-field approximation and the population's analysis of the bound state, it is found that these photoelectron peaks originate from the ionization of the excited state and the oscillations of these peaks are due to the superposition of their peak energy positions with the sideband energy positions. Furthermore, it is demonstrated that the energy positions of the maximum intensity of the photoelectron emission spectra move towards the higher energy end as the duration of the driving laser pulse extends. This phenomenon can be attributed to the fact that the main moment of ionization of atoms changes with the increasing duration of the driving laser pulse, thus allowing the real-time ionization of atoms to be probed using photoelectron emission spectra.

Minimum structure of high-order harmonic spectrum from molecular multi-orbital effects involving inner-shell orbitals.

The spectral features of high-order harmonic spectra can provide rich information for probing the structure and dynamics of molecules in intense laser fields. We theoretically study the high harmonic spectrum with the laser polarization direction perpendicular to the N2O molecule and find a minimum structure in the plateau region of the harmonic spectrum. Through analyzing the time-dependent survival probability of different electronic orbitals and the time-dependent wave packet evolution, it is found that this minimum position is caused by the harmonic interference of HOMO a, HOMO-1, and HOMO-3 a orbitals. Moreover, this interference minimum is discovered over a wide frequency range of 0.087 a.u. to 0.093 a.u., as well as a range of driving laser intensities with peak amplitudes between 0.056 a.u. and 0.059 a.u.. This study sheds light on the multi-electron effects and ultrafast dynamics of inner-shell electrons in intense laser pulses, which are crucial for understanding and controlling chemical reactions in molecules.

Enhancing harmonic brightness near the cutoff region by using laser pulses with a small positive chirp.

Efficient enhancement of harmonic brightness near the cutoff region is achieved by employing laser pulses with a small positive chirp in theory, where the laser intensity and frequency near the peak of the laser pulse are almost unchanged relative to the chirp-free field. The improvement of harmonic brightness is achieved under the condition that the ionization probability is almost unchanged. Through the analysis of the harmonics contributed by the rising and falling parts of the laser pulse, we have uncovered a "frequency compensation" mechanism that leads to an enhanced harmonic brightness near the cutoff region. Under appropriate chirp parameters, the harmonics contributed by the rising and falling parts can be constructively interfered in a smaller frequency range with greater intensity, thereby obtaining harmonics with good monochromaticity and high brightness. This study explains the mechanism of harmonic brightness enhancement from a new perspective, and provides a new idea for harmonic regulation without changing the ionization.

Circularly polarized attosecond light generation from OCS molecules irradiated by the combination of linear polarized infrared and orthogonal terahertz fields.

Researching ultrafast dynamics and creating coherent light sources will both benefit significantly from the establishment of polarization control in high-order harmonic generation (HHG). By employing the time-dependent density functional theory method, we investigate HHG of carbonyl sulfide molecules using a combination of a linear polarized infrared (IR) laser and a weaker orthogonal Terahertz (THz) field. Our findings show that by adjusting the amplitude of the THz field, the movement scale of electrons in the THz direction can be tuned, thereby one can control the harmonic intensity in the IR laser direction. This method allows for the creation of near-circularly polarized attosecond pulses. Furthermore, the ellipticity of the attosecond pulse may be changed by modifying the carrier-envelope phase of the IR laser pulse.

Influence of multiphoton resonance excitation on the above-threshold ionization of a hydrogen atom.

The photo-electron emission of a hydrogen atom irradiated by an ultraviolet laser pulse is investigated by numerically solving the time-dependent Schrödinger equation in momentum space. A subpeak structure with high intensity is observed in the photo-electron emission spectrum, and the peak of the enhanced structure shifts to a higher energy as the laser intensity increases. Through the strong-field approximation and the analysis of the population of the bound state , it is found that this subpeak structure is generated from the interference between the ionized electrons from the ground state and the ionized electrons from the 2p state after the resonant transition from the ground state to the 2p state. Analyzing the change rule of the photo-electron emission spectrum can further deepen the understanding of the energy change of the dressed bound state for an atom irradiated by an intense laser pulse.

Explanation of the anomalous redshift on a nonlinear X-ray Compton scattering spectrum by a bound electron.

Nonlinear Compton scattering is an inelastic scattering process where a photon is emitted due to the interaction between an electron and an intense laser field. With the development of X-ray free-electron lasers, the intensity of X-ray laser is greatly enhanced, and the signal from X-ray nonlinear Compton scattering is no longer weak. Although the nonlinear Compton scattering by an initially free electron has been thoroughly investigated, the mechanism of nonrelativistic nonlinear Compton scattering of X-ray photons by bound electrons is unclear yet. Here, we present a frequency-domain formulation based on the nonperturbative quantum electrodynamics to study nonlinear Compton scattering of two photons by an atom in a strong X-ray laser field. In contrast to previous theoretical works, our results clearly reveal the existence of a redshift phenomenon observed experimentally by Fuchs et al.(Nat. Phys.)11, 964(2015) and suggest its origin as the binding energy of the electron as well as the momentum transfer from incident photons to the electron during the scattering process. Our work builds a bridge between intense-laser atomic physics and Compton scattering processes that can be used to study atomic structure and dynamics at high laser intensities.

All-optical reconstruction of three-band transition dipole moments by the crystal harmonic spectrum from a two-color laser pulse.

When a bulk solid is irradiated by an intense laser pulse, transition dipole moments (TDMs) between different energy bands have an important influence on the ultra-fast dynamic process. In this paper, we propose a new all-optical method to reconstruct the k-dependent TDMs between multi-bands using a crystal high-order harmonic generation (HHG). Taking advantage of an obvious separation of bandgaps between three energy bands of an MgO crystal along the <001 > direction, a continuous harmonic spectrum with two plateaus can be generated by a two-color laser pulse. Furthermore, the first harmonic platform is mainly dominated by the polarization between the first conduction band and the valence band, and the second one is largely attributed to the interband HHG from the second conduction band and the valence band. Therefore, the harmonic spectrum from a single quantum trajectory can be adopted to map TDMs between the first, second conduction bands, and the valence one. Our work is of great significance for understanding the instantaneous properties of solid materials in the strong laser field, and will strongly promote the development of the HHG detection technology.

Calculation of high-order harmonic generation of atoms and molecules by combining time series prediction and neural networks.

High-order harmonic generation (HHG) from the interaction of ultra-intense laser pulses with atoms is an important tabletop short-wave coherent light source. Accurate quantum simulations of it present large computational difficulties due to multi-electron multidimensional effects. In this paper, the time-dependent response of hydrogen atoms is calculated using a time-series prediction scheme, the HHG spectrum is reconstructed very accurately. The accuracy of the forecasting is further improved by using a neural network scheme. This scheme is also applied to the simulation of the harmonic emission on multi-electron systems, and the applicability of the scheme is confirmed by the harmonic calculation of complex systems. This method is expected to simulate the nonlinear dynamic process of multi-electron atoms and molecules irradiated by intense laser pulses quickly and accurately.

Bioinspired Pd-Cu Alloy Nanoparticles as Accept Agent for Dye Degradation Performances.

Dye degradation is a key reaction in organic decomposition production through electron donor transferring. Palladium (Pd) is the best-known element for synthesis Pd-based catalyst, the surface status determines the scope of relative applications. Here we first prepare Pd-Cu alloy nanoparticles (NPs) by co-reduction of Cu(acac)2 (acac = acetylacetonate) and Pd(C5HF6O2)2 in the presence of sodium borohydride (NaBH4) and glutathione (GSH). The obtained Pd-Cu is about ~10 nm with super-hydrophilicity in aqueous mediums. The structural analysis clearly demonstrated the uniform distribution of Pd and Cu element. The colloidal solution keeps stability even during 30 days. Bimetallic Pd-Cu NPs shows biocompatibility in form of cell lines (IMEF, HACAT, and 239 T) exposed to colloidal solution (50 µg mL-1) for 2 days. It shows the catalytic multi-performance for dye degradation such as methyl orange (MO), rhodamine B (RhB), and methylene blue (MB), respectively. The as-synthesized nanoparticles showed one of the best multiple catalytic activities in the industrially important (electro)-catalytic reduction of 4-nitrophenol (4-NP) to corresponding amines with noticeable reduced reaction time and increased rate constant without the use of any large area support. In addition, it exhibits peroxidase-like activity in the 3, 3', 5, 5'-Tetramethylbenzidine (TMB) color test and exhibit obvious difference with previous individual metal materials. By treated with high intensity focused ultrasound filed (HIFU), Pd-Cu NPs might be recrystallized and decreased the diameters than before. The enhancement in catalytic performance is observed obviously. This work expedites rational design and synthesis of the high-hierarchy alloy catalyst for biological and environment-friendly agents.

Metal Cluster Triggered-Assembling Heterogeneous Au-Ag Nanoclusters with Highly Loading Performance and Biocompatible Capability.

In this work, we firstly report the preparation of heterogeneously assembled structures Au-Ag nanoclusters (NCs) as good drug carriers with high loading performance and biocompatible capability. As glutathione-protected Au and Ag clusters self-assembled into porous Au-Ag NCs, the size value is about 1.358 (±0.05) nm. The morphology characterization revealed that the diameter of Au-Ag NCs is approximately 120 nm, as well as the corresponding potential ability in loading performance of the metal cluster triggered-assembling process. Compared with individual components, the stability and loading performance of heterogeneous Au-Ag NCs were improved and exhibit that the relative biocompatibility was enhanced. The exact information about this is that cell viability was approximately to 98% when cells were incubated with 100 µg mL-1 particle solution for 3 days. The drug release of Adriamycin from Au-Ag NCs was carried out in PBS at pH = 7.4 and 5.8, respectively. By simulating in vivo and tumor microenvironment, the release efficiency could reach over 65% at pH = 5.8 but less than 30% at pH = 7.2. Using an ultrasound field as external environment can accelerate the assembling process while metal clusters triggered assembling Au-Ag NCs. The size and morphology of the assembled Au-Ag NCs can be controlled by using different power parameters (8 W, 13 W, 18 W) under ambient atmosphere. Overall, a novel approach is exhibited, which conveys assembling work for metal clusters triggers into heterogeneous structures with porous characteristic. Its existing properties such as water-solubility, stability, low toxicity and capsulation can be considered as dependable agents in various biomedical applications and drug carriers in immunotherapies.

New Insight into Assembled Fe3O4@PEI@Ag Structure as Acceptable Agent with Enzymatic and Photothermal Properties.

Metal-based enzyme mimics are considered to be acceptable agents in terms of their biomedical and biological properties; among them, iron oxides (Fe3O4) are treated as basement in fabricating heterogeneous composites through variable valency integrations. In this work, we have established a facile approach for constructing Fe3O4@Ag composite through assembling Fe3O4 and Ag together via polyethyleneimine ethylenediamine (PEI) linkages. The obtained Fe3O4@PEI@Ag structure conveys several hundred nanometers (~150 nm). The absorption peak at 652 nm is utilized for confirming the peroxidase-like activity of Fe3O4@PEI@Ag structure by catalyzing 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of H2O2. The Michaelis-Menten parameters (Km) of 1.192 mM and 0.302 mM show the higher catalytic activity and strong affinity toward H2O2 and TMB, respectively. The maximum velocity (Vmax) value of 1.299 × 10-7 M·s-1 and 1.163 × 10-7 M·s-1 confirm the efficiency of Fe3O4@PEI@Ag structure. The biocompatibility illustrates almost 100% cell viability. Being treated as one simple colorimetric sensor, it shows relative selectivity and sensitivity toward the detection of glucose based on glucose oxidase. By using indocyanine green (ICG) molecule as an additional factor, a remarkable temperature elevation is observed in Fe3O4@PEI@Ag@ICG with increments of 21.6 ∘C, and the absorption peak is nearby 870 nm. This implies that the multifunctional Fe3O4@PEI@Ag structure could be an alternative substrate for formatting acceptable agents in biomedicine and biotechnology with enzymatic and photothermal properties.

Xeno nucleic acid probes mediated methylation-specific PCR for single-base resolution analysis of N6-methyladenosine in RNAs.

Reliable and cost-effective quantification of RNA modifications at a specific gene locus is essential to elucidate the pathogenic mechanism encoded by RNA epigenetics. Current methods to quantify N6-methyladenosine (m6A) at specific sites can hardly satisfy the requirement of clinical application because epigenetic information is easily lost through polymerase chain reaction (PCR) assay or other isothermal amplification methods unless tedious pretreatment is applied. Herein, we propose a simple xeno nucleic acid (XNA) as a blocker probe to mediate the methylation specific reverse transcription quantitative polymerase chain reaction (MsRT-qPCR) assay to directly magnify the minor differences between epigenetic bases and unmodified bases in RNA. Strand displacement reactions selectively initiated between the reverse transcription primer (RT-primer) and the XNA probe at the m6A template given the affinity differences between the blocker probes and the m6A-modified RNA (m6A-RNA) and unmodified RNA (A-RNA). Thus, preferential amplification of m6A-RNA was allowed. Integration of a well-established oligo-modified Fe3O4@UiO-66-NH4 allowed purification of mRNA and lncRNA from cellular total RNA samples and greatly reduced the non-specific interference of m6A detection in real samples. Multiple specific sites of m6A in mRNA and lncRNA samples are also successfully quantified. The XNA probe-based m6A assay required only common and available lab equipment and materials, which can be applied in m6A-related fundamental studies and clinical diagnosis.

Proximity ligation assay mediated rolling circle amplification strategy for in situ amplified imaging of glycosylated PD-L1.

Glycosylated PD-L1 is a more reliable biomarker for immune checkpoint therapy and plays important roles in tumor immunity. Glycosylation of PD-L1 hinders antibody-based detection, which is partially responsible for the inconsistency between PD-L1 immunohistochemical results and therapeutic treatment response. Herein, we present a proximity ligation assay mediated rolling circle amplification (PLA-RCA) strategy for amplified imaging of glycosylated PD-L1 in situ. The strategy relies on a pair of DNA probes: an aptamer probe to specifically recognize cellular surface protein PD-L1 and a glycan conversion (GC) probe for metabolic glycan labeling. Upon proximity ligation of sequence binding to the two probes, the proximity ligation-triggered RCA occurs. The feasibility of the as-proposed strategy has been validated as it realized the visualization of PD-L1 glycosylation in different cancer cells and the monitoring of the variation of PD-L1 glycosylation during drug treatment. Thus, we envision the present work offers a useful alternative to track protein-specific glycosylation and potentially advances the investigation of the dynamic glycan state associated with the disease process.

Phage-Guided Targeting, Discriminative Imaging, and Synergistic Killing of Bacteria by AIE Bioconjugates.

New agents with particular specificity toward targeted bacteria and superefficacy in antibacterial activity are urgently needed in facing the crisis of worldwide antibiotic resistance. Herein, a novel strategy by equipping bacteriophage (PAP) with photodynamic inactivation (PDI)-active AIEgens (luminogens with aggregation-induced emission property) was presented to generate a type of AIE-PAP bioconjugate with superior capability for both targeted imaging and synergistic killing of certain species of bacteria. The targeting ability inherited from the bacteriophage enabled the bioconjugates to specifically recognize the host bacteria with preserved infection activity of phage itself. Meanwhile, the AIE characteristic empowered them a monitoring functionality, and the real-time tracking of their interactions with targets was therefore realized via convenient fluorescence imaging. More importantly, the PDI-active AIEgens could serve as powerful in situ photosensitizers producing high-efficiency reactive oxygen species (ROS) under white light irradiation. As a result, selective targeting and synergistic killing of both antibiotic-sensitive and multi-drug-resistant (MDR) bacteria were successfully achieved in in vitro and in vivo antibacterial tests with excellent biocompatibility. This novel AIE-phage integrated strategy would diversify the existing pool of antibacterial agents and inspire the development of promising drug candidates in the future.

Effect of interband polarization on a solid's high-order-harmonic generation just belowthe band gap.

A series of theoretical and experimental results has proved that harmonics below/above the band gap are produced mainly by the intraband current/interband polarization for solids in strong mid-infrared laser pulses. However, which mechanism dominates the harmonic process is still debated. In this work, based on simulating high-order-harmonic generation from an MgO crystal in a linearly polarized mid-infrared laser by solving semiconductor Bloch equations, we demonstrate that harmonics just below the band gap originate from the interference between intraband and interband currents. Furthermore, it is found that intensities of harmonics just below the band gap are apparently enhanced with an increase in the incident laser's strength. By analyzing the band dispersion and the transition dipole moment of the 001-cut MgO crystal, this can be attributed to the interband polarization between two conduction bands.

A fluorometric assay for rapid enrichment and determination of bacteria by using zirconium-metal organic frameworks as both capture surface and signal amplification tag.

A fluorometric assay was introduced to determine Acinetobacter baumannii (A. baumannii) in blood samples by utilizing Zr-MOFs both as functional coating for magnetic Fe3O4 nanoparticles to provide modification surface (Zr-mMOF) and as fluorescein carrier to produce fluorescence signals (F@UIO-66-NH2). Through strong Zr-O-P bonding, two distinct terminal phosphate-labeled A. baumannii and lipopolysaccharide (LPS) specific aptamers were attached onto Zr-MOFs to fabricate the magnetic core-shell capture probe (denoted as Zr-mMOF-p-Ab-Apt) and signal probe (denoted as F@UIO-66-NH2-p-LPS-Apt), respectively. After successive incubation with A. baumannii in blood samples and magnetic separation, the sandwich-type composite of capture probe/A. baumannii cells/signal probe was treated with high concentration of anionic phosphate ions to destroy the nano-structure of UIO-66-NH2 in the signal probe and fast release of fluorescein to produce amplified fluorescence signals. Due to the high aptamer modification efficiency of Zr-mMOF-p-Ab-Apt (up to 93%) and its strong affinity to A. baumannii, the enrichment efficiency of this capture probe has reached to 96.7%. Further, due to the high fluorescein loading efficiency of UIO-66-NH2 and our novel amplification strategy to destroy F@UIO-66-NH2-p-LPS-Apt to release and amplify fluorescein signals at 512 nm in the presence of high concentration of anionic phosphate ions, the sensitivity of this method has reached 10 cfu mL-1. This method allows enrichment and determination of A. baumannii within ~2.5 h. The limit of detection of A. baumannii in blood samples is 10 cfu mL-1 with a linear range of 101-105 cfu mL-1. This indicates the potential of this assay for diagnosis of bloodstream infection in early stage. Graphical abstractSchematic representation of sandwich-type fluorometric assay for Acinetobacter baumannii in blood samples with the capture probe (Zr-mMOF-p-Ab-Apt) and signal probe (F@UIO-66-NH2-p-LPS-Apt). The limit of detection is down to 10 cfu mL-1 with a linear range of 101-105 cfu mL-1.

Significantly enhanced conversion efficiency of high-order harmonic generation by introducing chirped laser pulses into scheme of spatially inhomogeneous field.

An effective scheme to enhance the yield of high-order harmonic generation originated from spatially inhomogeneous field through the interaction between few-cycle chirped laser pulses and a nano-tip structure is demonstrated. The conversion efficiency of harmonics from chirped laser pulses was significantly improved by nearly three more orders of magnitude than that of chirp-free pulses, and the cutoff energy of the corresponding harmonics was dramatically enhanced. By superimposing a series of properly selected orders of harmonics, isolated attosecond pulses of high intensity can be obtained. Furthermore, we compared the effects of different types of chirps on harmonics.