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基于Fosmid文库筛选山羊瘤胃微生物源蛋白酶基因及其表达验证

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西北农林科技大学学报（自然科学版） 2024年第4期52卷 28-38页

作者：杨凯尧刘功炜李林芳王智伟崔雯元杨雨鑫西北农林科技大学动物科技学院陕西杨凌712100

【目的】利用Fosmid文库功能筛选法,获得山羊瘤胃微生物源蛋白酶基因并进行原核表达。【方法】利用功能底物筛选法从山羊瘤胃微生物源Fosmid文库中筛选具有蛋白酶活性的克隆子,对其进行Illumina二代测序,对预测到的基因进行功能注释,根... 详细信息

【目的】利用Fosmid文库功能筛选法,获得山羊瘤胃微生物源蛋白酶基因并进行原核表达。【方法】利用功能底物筛选法从山羊瘤胃微生物源Fosmid文库中筛选具有蛋白酶活性的克隆子,对其进行Illumina二代测序,对预测到的基因进行功能注释,根据基因丰度结果确定后续研究的功能基因。利用大肠杆菌BL21(DE3)对功能基因进行诱导表达,对表达产物进行酶活性测定。通过CRISPR/Cas9基因编辑技术,将密码子优化后的功能基因分别敲入枯草芽孢杆菌C6基因组ctc位点并进行表达验证。【结果】从Fosmid文库1700个克隆子中筛选到1个具有蛋白酶活性的阳性克隆Pro4-C5。通过二代测序技术,鉴定出3个潜在的蛋白酶基因gene0833(内肽酶,EC编号:EC3.***.21.53)、gene0196(金属内肽酶,EC编号:EC3.4.24)和gene0585(羧肽酶,EC编号:EC3.***.17.14)以及1个L-天冬酰胺酶基因(gene0683,EC编号:EC3.***.1.1)。以pET-28a(+)为表达载体,通过大肠杆菌BL21(DE3)原核诱导表达发现,gene0585和gene0196未表达;gene0833表达的蛋白分子质量为87 ku,蛋白酶活性为10.45 U/mL;gene0683表达的蛋白分子质量为37 ku,L-天冬酰胺酶活性为88.52 U/mL,同时发现该蛋白也具有蛋白酶活性,酶活性为5.25 U/mL。将密码子优化后的gene0683和gene0833定点敲入枯草芽孢杆菌C6基因组中表达发现,gene0833未表达,gene0683表达的蛋白酶活性为109.72 U/mL,相比原始菌株(100.97 U/mL)显著提高了8.67%(P<0.01);L-天冬酰胺酶活性为31.63 U/mL,相比原始菌株(22.79 U/mL)显著提高了30.06%(P<0.01)。【结论】从山羊瘤胃微生物源Fosmid文库中筛选和鉴定出了2个具有蛋白酶活性的功能基因(gene0833和gene0683),均来源于微小杆菌属(Exiguobacterium),其中gene0683在大肠杆菌和枯草芽孢杆菌中的表达产物具有较高的蛋白酶和L-天冬酰胺酶酶活性。

关键词： Fosmid文库蛋白酶 L-天冬酰胺酶原核表达基因筛选菌株生产

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Defects and morphology engineering for constructing V_(s)-Ni_(3)S_(2)@V_(s)-Cu_(2)S nanotube heterojunction arrays toward efficient bifunctional electrocatalyst for overall water splitting

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Journal of Energy Chemistry 2024年第7期94卷 29-40页

作者： wenyuan Sun Alan Meng Lei Wang Guicun Li Jinfeng cui Yongkai Sun Zhenjiang Li College of Materials Science and Engineering Qingdao University of Science and TechnologyQingdao 266042ShandongChina Key Laboratory of Optic-electric Sensing and Analytical Chemistryfor Life Science MOE College of Chemistry and Molecular EngineeringQingdao Universityof Science and TechnologyQingdao 266042ShandongChina College of Environment and Safety Engineering Qingdao University of Science and TechnologyQingdao 266042ShandongChina Qingdao Huanghai University Qingdao 266000ShandongChina College of Electromechanical Engineering Qingdao University of Science and TechnologyQingdao 266061ShandongChina

The development of highly active,stable and inexpensive electrocatalysts for hydrogen production by defects and morphology engineering remains a great ***,S vacancies-rich Ni_(3)S_(2)@Cu_(2)S nan-otube heterojunction arrays were in-situ grown on copper foam(V_(s)-Ni_(3)S_(2)@V_(s)-Cu_(2)S NHAs/CF)for efficient electrocatalytic overall water *** the merits of nanotube arrays and efficient electronic mod-ulation drived by the OD vacancy defect and 2D heterojunction defect,the resultant V_(s)-Ni_(3)S_(2)@V_(s)-Cu_(2)S NHAs/CF electrocatalyst exhibits excellent electrocatalytic activity with a low overpotential of 47 mV for the hydrogen evolution reaction(HER)at 10 mA cm^(-2) current density,and 263 mV for the oxygen evolution reaction(OER)at 50 mA cm^(-2) current density,as well as a cell voltage of 1.48 V at 10 mA cm^(-2).Moreover,the nanotube heterojunction arrays endows V_(s)-Ni_(3)S_(2)@V_(s)-Cu_(2)S NHAs/CF with outstanding stability in long-term catalytic processes,as confirmed by the continuous chronopotentiom-etry tests at current densities of 10 mA cm^(-2) for 100 h.

关键词： Nanotubearrays Heterojunction Vacancy Bifunctional electrocatalyst Overall water splitting

Investigating the abundance enrichment pattern of heavy elements in the only observed CEMP-r/s star J004441.04-732136.4 of the SMC

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Science China(Physics,Mechanics & Astronomy) 2014年第6期57卷 1201-1205页

作者： cui wenyuan ZHANG Bo ZHAO Gang School of Space Science and Physics Shandong University at Weihai College of Physics Science and Information Engineering Hebei Normal University National Astronomical Observatories Chinese Academy of Sciences

The post-AGB star J004441 is the first and the only one CEMP-r/s star found in ***,we investigate the observed abundance pattern of the heavy elements using our parametric model.A consistent fitting results was obtained for the sample *** on the low r=0.08,the s-process nucleosynthesis occurred in the interior is supposed to belong to the single neutron-exposure *** median value ofτ0=0.44(T90.348)1/2mbarn-1supports a higher efficiency of the s-process nucleosynthesis relative to J004441 than that of the solar system,however,the value is not sufficiently high to favor the formation of a lead ***,J004441does not belong to lead star *** large Csvalue of J004441 supports the intrinsic characteristic of the *** Crvalue is similar with that found in halo CEMP-r/s stars,which indicates that the r-process contributions is critical during heavy element *** star has a metallicity of[Fe/H]=-1.34,which is larger than that of Galaxy halo CEMP-r/s *** reason may be because of the different history of metallicity enrichment between the SMC and the Galaxy halo.

关键词：富集规律重元素明星金属丰度 S-过程核合成 AGB星参数模型

Soft initial-rotation and H_∞ robust constant rotational speed control for rotational MEMS gyro

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Journal of Systems Engineering and Electronics 2009年第3期20卷 583-589页

作者： Ma Gaoyin Chen wenyuan cui Feng Zhang Weiping Wang Liqi National Key Lab. of Nano/Micro Fabrication Technology Key Lab. for Thin Film and Microfabrication of Ministry of Education Inst. of Micro and Nano Science and Technology Shanghai Jiaotong Univ.Shanghai 200240 P. R. China

A novel soft initiai-rotation control system and an H∞ robust constant rotational speed controller （RCRSC） for a rotational MEMS （micro-electro-mechanical system） gyro are presented. The soft initial-rotation control system can prevent the possible tumbling down of the suspended rotor and ensure a smooth and fast initial-rotation process. After the initial-rotation process, in order to maintain the rotational speed accurately constant, the RCRSC is acquired through the mixed sensitivity design approach. Simulation results show that the actuation voltage disturbances from the internal carrier waves in the gyro is reduced by more than 15.3 dB, and the speed fluctuations due to typical external vibrations ranging from 10 Hz to 200 Hz can also be restricted to 10^-3 rad/s order.

关键词： micro-electro-mechanicai system gyro rotation mixed sensitivity design H∞ robust controller

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面向中国式现代化的税制改革研究与探索

税务研究 2023年第9期 101-107页

作者：崔文苑国家税务总局税收宣传中心

进入新时代以来,中国税制改革取得了系列伟大成就。面向实现中国式现代化的宏伟目标,必须深入贯彻落实党的二十大关于税制改革的决策部署,立足新发展阶段,贯彻新发展理念,围绕构建新发展格局、实现高质量发展,按照《中华人民共和国国民... 详细信息

进入新时代以来,中国税制改革取得了系列伟大成就。面向实现中国式现代化的宏伟目标,必须深入贯彻落实党的二十大关于税制改革的决策部署,立足新发展阶段,贯彻新发展理念,围绕构建新发展格局、实现高质量发展,按照《中华人民共和国国民经济和社会发展第十四个五年规划和2035年远景目标纲要》要求,坚持税收法定原则、公平与效率相统一原则、宏观税负稳定原则,从优化税制结构、提高税制科学性,逐步降低名义税率、提高税制竞争力,健全地方税体系、保障地方财政收入三个方面入手,深入推进税制改革,进一步发挥税收在国家治理中的基础性、支柱性、保障性作用。

关键词：税制改革中国式现代化税制结构名义税率地方税体系

The fluid inclusions in jadeitite from Pharkant area, Myanmar

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Chinese Science Bulletin 2000年第20期45卷 1896-1901,1923页

作者： SHI Guanghai cui wenyuan WANG Changqiu ZHANG Wenhuai Department of Geology Peking UniversityBeijing 100871China Institute of Resource China University of GeosciencesWuhan 430074China

A lot of liquid-gas and liquid-gas-solid inclusions were found in Pharkant jadeitites, northwestern Myanmar and their characteristics, geological setting and porphyroclastic jadeites with inclusions in them were described in detail. The results analyzed by Raman spectrometer showed that the component of liquid-gas phase and solid phase (daughter minerals) in fluid inclusions is H2O + CH4 and jadeite separately. The results indicated that Pharkant jadeitites were crystallized from H2O + CH4 bearing jadeitic melt which may originate from mantle. The P-T conditions in which the jadeitites were crystallized were speculated to be T>650℃, P>1.5 GPa.

关键词： Pharkant Myanmar jadeitite jadeite fluid inclusion H2O + CH4 jadeitic melt mantle.

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用光影讲好中国税收故事的实践探索

电影评介 2023年第22期 94-96页

作者：崔文苑国家税务总局税收宣传中心宣传处

近年来,税务部门顺应融媒体发展趋势,以大学生电影节“中国税收”特别单元等赛事为契机,不断探索短视频创作新路径,创作了丰富的税收短视频作品。本文结合当下税收短视频创作的现状和难点,探索如何进一步抓好税收短视频创作,用光影讲好... 详细信息

近年来,税务部门顺应融媒体发展趋势,以大学生电影节“中国税收”特别单元等赛事为契机,不断探索短视频创作新路径,创作了丰富的税收短视频作品。本文结合当下税收短视频创作的现状和难点,探索如何进一步抓好税收短视频创作,用光影讲好中国税收故事,持续扩大税收文化的覆盖面和影响力。

关键词：税收文化视频创作税务部门短视频实践探索光影现状和难点覆盖面

辽西建平变质杂岩中花岗岩类岩石的锆石蒸发年龄及其地质意义

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北京大学学报（自然科学版） 1999年第4期35卷 542-549页

作者：王长秋崔文元北京大学地质学系

建平变质杂岩是辽宁西都太古代最下部地质单元，是华北地台北缘麻粒岩相带的重要组成部分。它由少量连续性很差的表壳岩及大量的ＴＴＧ岩石和其他深成侵入体组成。采用单颗粒锆石Ｐｂ蒸发法对其中的ＴＴＧ岩石及其他侵入岩进行了年龄测... 详细信息

建平变质杂岩是辽宁西都太古代最下部地质单元，是华北地台北缘麻粒岩相带的重要组成部分。它由少量连续性很差的表壳岩及大量的ＴＴＧ岩石和其他深成侵入体组成。采用单颗粒锆石Ｐｂ蒸发法对其中的ＴＴＧ岩石及其他侵入岩进行了年龄测定。结果表明：本区ＴＴＧ花岗质岩浆约于２５２１Ｍａ开始活动，并于２５００Ｍａ左右达到高峰，形成了大量的ＴＴＧ质侵入体（包括部分喷出岩）；紧随其后，本区发生强烈的高级变质变形事件，此事件约于２４８３～２４９０Ｍａ达到高峰，使得已形成的岩石变成麻粒岩、紫苏花岗岩及斜长紫苏花岗岩等；于２４７２Ｍａ就位的与退变质作用有关的后构造花岗岩标志着本区高级变质变形事件的结束。据此，可以推测建平变质杂岩可能形成于陆块形成后不久即发生大陆碰撞和地壳加厚的晚太古活动大陆边缘的环境中。

关键词：锆石年龄 TTG岩石变质杂岩花岗岩铅蒸发法

Insights into varicella-zoster virus assembly from the B-and C-capsid at near-atomic resolution structures

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hLife 2024年第2期2卷 64-74页

作者： Lei Cao Nan Wang Zhe Lv wenyuan Chen Zhonghao Chen Lifei Song Xueyan Sha Guiqiang Wang Yaling Hu Xiaojun Lian Guoliang cui Jinyan Fan Yaru Quan Hongrong Liu Hai Hou Xiangxi Wang CAS Key Laboratory of Infection and Immunity National Laboratory of MacromoleculesInstitute of BiophysicsChinese Academy of SciencesBeijingChina Key Laboratory for Matter Microstructure and Function of Hunan Province Key Laboratory of Low-dimensional Quantum Structures and Quantum ControlSchool of Physics and ElectronicsHunan Normal UniversityHunanChina Institute of Medical Research Northwestern Polytechnical UniversityShanxiChina Sinovac(Dalian)Vaccine Technology Co. Ltd.LiaoningChina Sinovac Biotech Ltd. BeijingChina National Institutes for Food and Drug Control BeijingChina

Varicella-zoster is a highly communicable virus that can be transmitted through the airborne *** one quarter of people are infected with this *** studies have described the structure of A-capsid and a blurred reconstruction of the C-capsid with icosahedral *** this study,we have determined the more precise detailed structures of the varicella-zoster virus(VZV)B-and C-capsid in icosahedral symmetry using a combination of block-based reconstruction and symmetry relaxation *** addition,we are reporting structural details of the portal vertex reconstructions in five-fold symmetry and portal reconstructions in twelve-fold *** structures unveil the basis for the high thermal stability of the VZV *** conformational flexibility of structural elements of the capsid plays a role in the assembly of the capsid and drives processes critical for the viral life *** results of the study open up new avenues for the development of drugs against a highly prevalent and contagious pathogen.

关键词： varicella-zoster virus capsid structure virus assembly