一种具有高β-葡聚糖酶活性的几丁质外切酶基因的克隆、表达和性质鉴定

【目的】表达并鉴定来源于维氏气单胞菌的几丁质酶Chi92并研究其作为水产饲用酶的有效性。【方法】自A.veronii B565中克隆chi92基因并在Pichia pastoris GS115中进行表达,对表达成功的Chi92进行分离纯化和生化鉴定。最后将Chi92添加到含有毕赤酵母粉的饲料中饲喂斑马鱼2周,研究Chi92添加对斑马鱼生长、饲料利用率、肠道微绒毛形态和抗病性能的影响。【结果】chi92基因编码具有864个氨基酸残基的多肽。Chi92在p H 6.0和40°C时表现最佳酶活。Chi92对蛋白酶有抗性,同时酶活不受金属离子显著影响。Chi92具备高几丁质酶活(69.4 U/m L)。以胶体几丁质和β-1,3-1,4-葡聚糖作为底物时,比活力分别为809.2 U/mg和235.6 U/mg。薄层层析和电喷雾电离质谱联用技术均表明N-乙酰葡糖胺二聚体是Chi92酶解胶体几丁质的主要产物。Chi92在对酵母细胞壁的降解方面比其他几丁质酶性能更加优良。经过2周饲喂,添加有Chi92的饲料显著提高了斑马鱼肠道微绒毛的高度和密度,同时斑马鱼的生长,饲料利用率,以及抗病性能均得到了一定提高。【结论】Chi92具有p H稳定性、抗逆性和高酵母细胞壁降解功能,能较好地作为饲用酶用于温水水产养殖。     

Polymerase chain reaction–hybridization method using urease gene sequences for high-throughput Ureaplasma urealyticum and Ureaplasma parvum detection and differentiation

In this article, we discuss the polymerase chain reaction (PCR)–hybridization assay that we developed for high-throughput simultaneous detection and differentiation of Ureaplasma urealyticum and Ureaplasma parvum using one set of primers and two specific DNA probes based on urease gene nucleotide sequence differences. First, U. urealyticum and U. parvum DNA samples were specifically amplified using one set of biotin-labeled primers. Furthermore, amine-modified DNA probes, which can specifically react with U. urealyticum or U. parvum DNA, were covalently immobilized to a DNA–BIND plate surface. The plate was then incubated with the PCR products to facilitate sequence-specific DNA binding. Horseradish peroxidase–streptavidin conjugation and a colorimetric assay were used. Based on the results, the PCR–hybridization assay we developed can specifically differentiate U. urealyticum and U. parvum with high sensitivity (95%) compared with cultivation (72.5%). Hence, this study demonstrates a new method for high-throughput simultaneous differentiation and detection of U. urealyticum and U. parvum with high sensitivity. Based on these observations, the PCR–hybridization assay developed in this study is ideal for detecting and discriminating U. urealyticum and U. parvum in clinical applications.     

Deletion of pancreatic β‐cell adenosine kinase improves glucose homeostasis in young mice and ameliorates streptozotocin‐induced hyperglycaemia

Severe reduction in the β‐cell number (collectively known as the β‐cell mass) contributes to the development of both type 1 and type 2 diabetes. Recent pharmacological studies have suggested that increased pancreatic β‐cell proliferation could be due to specific inhibition of adenosine kinase (ADK). However, genetic evidence for the function of pancreatic β‐cell ADK under physiological conditions or in a pathological context is still lacking. In this study, we crossed mice carrying LoxP‐flanked Adk gene with Ins2‐Cre mice to acquire pancreatic β ‐cell ADK deficiency (Ins2‐Cre^±Adk^fl/fl) mice. Our results revealed that Ins2‐Cre^+/‐Adk^fl/fl mice showed improved glucose metabolism and β‐cell mass in younger mice, but showed normal activity in adult mice. Moreover, Ins2‐Cre^±Adk^fl/fl mice were more resistant to streptozotocin (STZ) induced hyperglycaemia and pancreatic β‐cell damage in adult mice. In conclusion, we found that ADK negatively regulates β‐cell replication in young mice as well as under pathological conditions, such as STZ induced pancreatic β‐cell damage. Our study provided genetic evidence that specific inhibition of pancreatic β‐cell ADK has potential for anti‐diabetic therapy.     

Steric Engineering of Alkylthiolation Side Chains to Finely Tune Miscibility in Nonfullerene Polymer Solar Cells

Morphology and miscibility control are still a great challenge in polymer solar cells. Despite physical tools being applied, chemical strategies are still limited and complex. To finely tune blend miscibility to obtain optimized morphology, chemical steric engineering is proposed to systemically investigate its effects on optical and electronic properties, especially on a balance between crystallinity and miscibility. By changing the alkylthiol side chain orientation different steric effects are realized in three different polymers. Surprisingly, the photovoltaic device of the polymer PTBB‐m with middle steric structure affords a better power conversion efficiency, over 12%, compared to those of the polymers PTBB‐o and PTBB‐p with large or small steric structures, which could be attributed to a more balanced blend miscibility without sacrificing charge‐carrier transport. Space charge‐limited current, atomic force microscopy, grazing incidence wide angle X‐ray scattering, and resonant soft X‐ray scattering measurements show that the steric engineering of alkylthiol side chains can have significant impacts on polymer aggregation properties, blend miscibility, and photovoltaic performances. More important, the control of miscibility via the simple chemical approach has preliminarily proved its great potential and will pave a new avenue for optimizing the blend morphology.     

Radially Oriented Single‐Crystal Primary Nanosheets Enable Ultrahigh Rate and Cycling Properties of LiNi0.8Co0.1Mn0.1O2 Cathode Material for Lithium‐Ion Batteries

Ni‐rich Li[Ni_xCo_yMn_1?x?y]O₂ (x ≥ 0.8) layered oxides are the most promising cathode materials for lithium‐ion batteries due to their high reversible capacity of over 200 mAh g^?1. Unfortunately, the anisotropic properties associated with the α‐NaFeO₂ structured crystal grains result in poor rate capability and insufficient cycle life. To address these issues, a micrometer‐sized Ni‐rich LiNi_0.8Co_0.1Mn_0.1O₂ secondary cathode material consisting of radially aligned single‐crystal primary particles is proposed and synthesized. Concomitant with this unique crystallographic texture, all the exposed surfaces are active {010} facets, and 3D Li⁺ ion diffusion channels penetrate straightforwardly from surface to center, remarkably improving the Li⁺ diffusion coefficient. Moreover, coordinated charge–discharge volume change upon cycling is achieved by the consistent crystal orientation, significantly alleviating the volume‐change‐induced intergrain stress. Accordingly, this material delivers superior reversible capacity (203.4 mAh g^?1 at 3.0–4.3 V) and rate capability (152.7 mAh g^?1 at a current density of 1000 mA g^?1). Further, this structure demonstrates excellent cycling stability without any degradation after 300 cycles. The anisotropic morphology modulation provides a simple, efficient, and scalable way to boost the performance and applicability of Ni‐rich layered oxide cathode materials.     

Rational Design of Hierarchical “Ceramic‐in‐Polymer” and “Polymer‐in‐Ceramic” Electrolytes for Dendrite‐Free Solid‐State Batteries

Solid polymer electrolytes as one of the promising solid‐state electrolytes have received extensive attention due to their excellent flexibility. However, the issues of lithium (Li) dendrite growth still hinder their practical applications in solid‐state batteries (SSBs). Herein, composite electrolytes from “ceramic‐in‐polymer” (CIP) to “polymer‐in‐ceramic” (PIC) with different sizes of garnet particles are investigated for their effectiveness in dendrite suppression. While the CIP electrolyte with 20 vol% 200 nm Li_6.4La₃Zr_1.4Ta_0.6O₁₂ (LLZTO) particles (CIP‐200 nm) exhibits the highest ionic conductivity of 1.6 × 10^?4 S cm^?1 at 30 °C and excellent flexibility, the PIC electrolyte with 80 vol% 5 µm LLZTO (PIC‐5 µm) shows the highest tensile strength of 12.7 MPa. A sandwich‐type composite electrolyte (SCE) with hierarchical garnet particles (a PIC‐5 µm interlayer sandwiched between two CIP‐200 nm thin layers) is constructed to simultaneously achieve dendrite suppression and excellent interfacial contact with Li metal. The SCE enables highly stable Li plating/stripping cycling for over 400 h at 0.2 mA cm^?2 at 30 °C. The LiFePO₄/SCE/Li cells also demonstrate excellent cycle performance at room temperature. Fabricating sandwich‐type composite electrolytes with hierarchical filler designs can be an effective strategy to achieve dendrite‐free SSBs with high performance and high safety at room temperature.     

Introgression lines of Solanum sitiens,a wild nightshade of the Atacama Desert,in the genome of cultivated tomato

The wild tomato relative Solanum sitiens is a xerophyte endemic to the Atacama Desert of Chile and a potential source of genes for tolerance to drought, salinity and low‐temperature stresses. However, until recently, strong breeding barriers prevented its hybridization and introgression with cultivated tomato, Solanum lycopersicum L. We overcame these barriers using embryo rescue, bridging lines and allopolyploid hybrids, and synthesized a library of introgression lines (ILs) that captures the genome of S. sitiens in the background of cultivated tomato. The IL library consists of 56 overlapping introgressions that together represent about 93% of the S. sitiens genome: 65% in homozygous and 28% in heterozygous (segregating) ILs. The breakpoints of each segment and the gaps in genome coverage were mapped by single nucleotide polymorphism (SNP) genotyping using the SolCAP SNP array. Marker‐assisted selection was used to backcross selected introgressions into tomato, to recover a uniform genetic background, to isolate recombinant sub‐lines with shorter introgressions and to select homozygous genotypes. Each IL contains a single S. sitiens chromosome segment, defined by markers, in the genetic background of cv. NC 84173, a fresh market inbred line. Large differences were observed between the lines for both qualitative and quantitative morphological traits, suggesting that the ILs contain highly divergent allelic variation. Several loci contributing to unilateral incompatibility or hybrid necrosis were mapped with the lines. This IL population will facilitate studies of the S. sitiens genome and expands the range of genetic variation available for tomato breeding and research.     

Eg5 high expression predicts dismal prognosis in epithelial ovarian cancer

The expression of kinesin spindle protein (Eg5) and its significance of clinical prognosis of patients with epithelial ovarian cancer (EOC) were evaluated in this study. Forty-five fresh frozen tissue samples for quantitative real-time polymerase chain reaction (qPCR) and 196 samples for immunohistochemistry (IHC) analysis with tissue microarray (TMA) were applied to characterize Eg5 mRNA and protein expressions in EOC. The correlation between clinical parameters and Eg5 protein expression was investigated using statistical analysis. The expression of Eg5 protein was significantly higher in EOC tissues compared with that in corresponding non-cancerous tissues (P < 0.05). The high Eg5 expression was significantly correlated with older age (P = 0.003), higher stage (P = 0.001), presence of metastasis (P = 0.041) and higher CA125 serum level (P = 0.013). For univariate analysis, associated prognostic markers in patients with ovarian cancer were analyzed for correlations with poor overall survival, including Eg5 (P = 0.011), age (P = 0.001), FIGO stage (P = 0.011), CA125 serum level (P = 0.001), lymph nodes (P = 0.012), and metastasis (P = 0.001). For multivariate analysis, Eg5 expression, FIGO stage and age were independent factors found contributing to a largely unfavorable prognosis in patients with ovarian cancer. In conclusion, the high expression of Eg5 is correlated with an unfavorable prognosis in EOC.