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PCR-based site-specific mutagenesis of peptide antibiotics FALL-3. and its biologic activities

Acta Pharmacologica Sinica 2004年第2期25卷 113-119页

作者： Yun-xia YANG2,3. Yun FENG3. Bo-yao WANG3.Qi WU3.2Department of Pharmacology,3.esearch Unit of Infection & Immunity, West-china school of Preclinical and Forensic medicine, Sichuan University, Chengdu 610041, china Department of Pharmacology West-China School of Preclinical and Forensic Medicine Sichuan University Chengdu 610041 Research Unit of Infection & Immunity West-China School of Preclinical and Forensic Medicine Sichuan University Chengdu 610041

AIM: To construct PGEX-1λT-FALL-3. expression vector and its mutant vector, and study the relationship of function and structure. METHODS: A cDNA encoding mature FALL-3. was cloned from SPCA-1 cell mRNA and the prokaryotic expression vector PGEX-1λT-FALL-3. was constructed. Two kinds of polymerase chain reaction (PCR) for the site-direction mutagenesis were used to construct FALL-3. mutant expression vector, FALL-3.-Lys- 3. and FALL-3.-Lys-24. Minimal effective concentration, minimal inhibitory concentration, and minimal bacteri- cidal concentration were used to assay the antibacterial activities of these peptides. Effects of different solution on the antibacterial activity of FALL-3. and FALL-3.-Lys-3. were observed by CFU determination. The hemolytic effects of these peptides were also examined on human red blood cells. RESULTS: Two site-specific mutants FALL-3.-Lys-3. and FALL-3.-Lys24 were obtained by PCR-induced mutagenesis. In comparison with two-step PCR which required two pairs of primers, one step PCR which required one pair of primers is a simple and efficient method for the PCR based site-specific mutagenesis. Using the prokaryotic expression system, the E coli-based products of recombinant FALL3. and its mutant peptides were also obtained. The antibacterial assay showed that FALL-3.-Lys-3. and FALL-3.-Lys24 were more potential in the antibacterial activity against E coli ML3.p and Pseudomonas aeruginosa ATCC27853.than that of FALL-3., and no increase in hemolysis was observed at the antibacterial concentrations. The antibacterial activity of FALL-3.-Lys-3. against E coli was more potent than that of FALL-3. in NaCl-containing LB medium, while its activity was almost the same as FALL-3. in SO4 containing 2- Medium E. CONCLUSION: PCR-based mutagensis is a useful model system for studying the structure and function relationship of antimicrobial peptides. Keeping α-helical conformation of FALL-3. and increasing net positive charge can increase the antibacter

关键词： FALL 3. peptide antibiotics polymerase chain reaction mutation hemolysis

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Stable Skin-specific Overexpression of Human CTLA4-Ig in Transgenic Mice through Seven Generations

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Acta Biochimica et Biophysica Sinica 2006年第3期38卷 171-178页

作者： Yong WANG~1 Yong NI~(1,3. Hong WEI~1* Feng-Chao WANG~2 Liang-Peng GE~1 Xiang GAO~41 Department of Animal Science,College of basic medicine,Third Military Medical University,Chongqing 40003.,china 2 Institute of Combined Injury,College of Military Preventive medicine,Third Military Medical University,Chongqing 40003.,china 3.school of Life Sciences,Southwest University,Chongqing 400715,china 4 Animal Model Research Institute,Nanjing University,Nanjing 210093.china Department of Animal Science College of Basic Medicine Third Military Medical UniversityChongqing 400038 China school of Life Sciences Southwest UniversityChongqing 400715 China Institute of Combined Injury College of Military Preventive Medicine Third Military Medical UniversityChongqing 400038 China Animal Model Research Institute Nanjing UniversityNanjing 210093 China

Skin graft rejection is a typical cellular immune response,mainly mediated by T *** lymphocyte associated antigen 4-immunoglobin (CTLA4-Ig) extends graft survival by blocking the T cellco-stimulation pathway and inhibiting T cell *** investigate the efficacy of CTLA4-Ig in prolong-ing skin graft survival,human CTLA4-Ig (hCTLA4-Ig) was engineered to overexpress in mouse skin bytransgenesis using the K14 *** transcription-polymerase chain reaction (RT-PCR) and West-ern blot assay indicated that the expression of CTLA4-Ig remained skin-specific and relatively constantcompared to the internal control protein,AKT,through seven *** presence and concentrationof the hCTLA4-Ig protein in transgenic mouse sera was determined by enzyme-linked immunosorbent assay(ELISA),and the results indicated that the serum CTLA4-Ig concentration also remained constant *** of transgenic mouse skins grafted onto rat wounds was remarkably prolonged com-pared to that of wild-type skins from the same mouse strain,and remained comparable among all *** suggested that the bioactive hCTLA4-Ig protein was stably expressed in transgenical micethrough at least seven generations,which was consistent with the stable skin-specific CTLA4-Ig *** results demonstrated that the transgenic expression of hCTLA4-Ig in skin driven by the K14 promoterremained constant through generations,and a transgenic line can be established to provide transgenic skinwith extended survival reproducibly.

关键词： cytotoxic T lymphocyte antigen 4-Ig transgenic mice skin K14 promoter

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Synthesis and anticonvulsant activity of 7-benzylamino-4,5-dihydro-[1,2,4] triazolo[4,3.a] quinolines

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Progress in Natural Science:Materials International 2007年第9期17卷 1104-1108页

作者： Cui Xiangshu1,2,Guan Liping3.Li Hailan3.Piao Huri3.and Quan Zheshan1,3.1.Key Laboratory of Organic Functional Factors of the Changbai Mountain,Ministry of Education,Yanbian University,Yanji 13.002,china 2.College of basic medicine,Yanbian University,Yanji 13.000,china 3.College of Pharmacy,Yanbian University,Yanji 13.000,china) Key Laboratory of Organic Functional Factors of the Changbai Mountain Ministry of Education Yanbian University Yanji 133002 China College of basic medicine Yanbian University Yanji 133000 China College of Pharmacy Yanbian University Yanji 133000 China

A series of 7-substituted-benzylamino-4,5-dihydro-[1,2,4]triazolo[4,3.a]quinoline derivatives was synthesized and evaluated for their anticonvulsant *** subcutaneous pentylenetetrazole test(sc-PTZ)demonstrated that the most effective compound in controlling the sc-PTZ induced seizure was 7-(3.bromine-benzylamino)-4,5-dihydro-[1,2,4]triazolo[4,3.a]quinoline(4j)with an ED50 of 5.0 mg/kg and the PI of 20.7,which was also safer than the reference *** the maximal electroshock test(MES)demonstrated that among these derivatives,7-(3.fluorobenzylamino)-4,5-dihydro-[1,2,4]trizolo[4,3.a]quinoline(4i),with an ED50 of 15.3.mg/kg and the PI of 7.2,was the safest in MES ***,their neurotoxicities were measured by the rotarod neurotoxicity test,and the results showed that all derivatives possessed lower neurotoxicity.

关键词： quinoline triazole synthesis anticonvulsant epilepsia

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AICAR inhibits proliferation and induced S-phase arrest,and promotes apoptosis in CaSki cells

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Acta Pharmacologica Sinica 2007年第12期28卷 1984-1990页

作者： Tong-ju GUAN~2,Feng-jin QIN~3.Jian-hai DU~4,Li Geng~3.You-yi ZHANG~(4,5),Min LI~(2,5)~2 Institute of Pathology,The school of basic medicine,Lanzhou University,Lanzhou 73.000,{3. ~3.Department of Gynaecology andObstetrics,Peking University Third Hospital,Beijing 100083.{3. ~4 Institute of Vascular medicine,Peking University Third Hospital andKey Laboratory of Molecular Cardiovascular Sciences Ministry of Education,Beijing 100083.{3. Institute of Pathology The School of Basic Medicine Lanzhou University Lanzhou China Department of Gynaecology and Obstetrics Peking University Third Hospital Beijing China Institute of Vascular medicine Peking University Third Hospital and Key Laboratory of Molecular Cardiovascular Sciences Ministry of Education Beijing China

Aim:The aim of the present study was to determine the effect of 5-aminoimidazole-4-carboxamide-ribonucleoside （AICAR） on proliferation,cell cycle,and apoptosisin the human epithelial cervical cancer cell line CaSki ***:Cell countand 3.（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide assay were usedto determine cell proliferation and *** 3.258 staining was con-ducted to distinguish the apoptotic *** cycle and Annexin-V/propidiumiodide staining were analyzed by fluorescence-activated cell sorting （FACS）.AWestern blot assay was used to evaluate the expression of AKT （also known asprotein kinase B）,mammalian target of rapamycin （mTOR）,p53.and extracellularsignal-regulated kinase （ERK）.Results:AICAR （500 μmol/L） significantly inhibi-ted the proliferation of CaSki cells treated for 24,48,and 72 h as determined by *** cells at the G1and G2phases were dramatically decreased while cells atthe S phase were increased in response to AICAR treatment for 24,48,and 72 *** MTT assay showed less viable cells and Hoechst fluorescent staining showedmore apoptotic cells upon AICAR *** results of the Annexin-Vstaining demonstrated a time-dependent increase of apoptosis in cells treatedwith AICAR for 24,3.,and 48 ***,AICAR activated caspase-3.in atime-dependent *** was also found that AICAR inhibited the phosphory-lation of AKT and mTOR,which are important kinases regulating cell growth *** stimulation obviously increased the expression of the tumorsuppressor p53.and the phosphorylation of ***:AICAR inhibitedproliferation and induced S phase arrest and promoted apoptosis in CaSki cells,which might be mediated by the downregulation of the AKT/mTOR pathway andthe upregulation of the p53.ERK pathway.

关键词： AICAR proliferation apoptosis CaSki cells

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Conceptual design,data model and key techniques of“3. Assistant”

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中国有色金属学会会刊：英文版 2011年第S3期21卷 692-698页

作者： LI Qing-yuan1, ZHANG Xuan2, JIN Yang2, ZHOU Hui3. WANG Qian-jin3.WEI Zhan-yin1 1. Key Laboratory of Geo-Informatics, Chinese Academy of Surveying and Mapping, Beijing 10083., china 2. school of Geo-Science and Surveying Engineering, china University of Mining and Technology, Beijing 100083. china 3. school of Geomatics, Liaoning Technical University, Fuxin 123.00, china Key Laboratory of Geo-Informatics Chinese Academy of Surveying and Mapping Beijing 100830 China school of Geo-Science and Surveying Engineering China University of Mining and Technology Beijing 100083 China school of Geomatics Liaoning Technical University Fuxin 123000 China

Conceptual design, data model and key techniques of "Geo-engineer 3. assistant", an underground 3. GIS oriented to geology and mine application, were discussed. In particular, the importance of high-order smoothing triangular pieces to surface's fine expression in geological modeling was emphasized. The contour mode is the most favorite mode of the three modes about surface expression: DEM, TIN and contour. Topology in underground 3. modeling with CAGD was compared. From the view of boundary and coboundary, which is soul of IS 19107 (Geographic information spatial schema), 3. underground topology was re-examined.

关键词： 3. assistant true 3. GIS fine expression of surface contour mode 3. topology

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传染病多点触发智慧化监测预警系统关键技术专家共识

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中华医学杂志 2024年第32期104卷 1-15页

作者：中国医学科学院北京协和医学院群医学及公共卫生学院北京大学公共卫生学院中国疾病预防控制中心传染病管理处四川大学华西公共卫生学院中国科学院地理科学与资源研究所首都医科大学附属北京地坛医院中华医学会公共卫生分会

有效的传染病监测预警系统是公共卫生安全体系的重要组成部分, 对预防与控制传染病暴发流行至关重要, 提升监测预警能力是推动疾病预防控制事业高质量发展的当务之急。专家组结合公共卫生与预防医学、临床医学、管理学、数据科学、计算... 详细信息

有效的传染病监测预警系统是公共卫生安全体系的重要组成部分, 对预防与控制传染病暴发流行至关重要, 提升监测预警能力是推动疾病预防控制事业高质量发展的当务之急。专家组结合公共卫生与预防医学、临床医学、管理学、数据科学、计算机科学等多学科专家的研究成果与实践经验, 经过反复多轮专家研讨, 形成了传染病多点触发智慧化监测预警系统关键技术专家共识。本共识内容主要包括传染病多点触发智慧化监测预警系统的相关概念和定义, 关键技术框架, 多渠道监测预警数据来源、获取与治理, 预警方法分类, 多点触发智慧化监测预警关键流程及路径, 多点触发预警及综合研判, 预警信号响应以及预警效果评价等方面, 以期为传染病多点触发智慧化监测预警系统建设及应用提供技术参考。

关键词：传染病监测预警多点触发智慧化

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Mining and Integrating Reliable Decision Rules for Imbalanced Cancer Gene Expression Data Sets

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Tsinghua Science and Technology 2012年第6期17卷 666-673页

作者： Hualong Yu 1 , Jun Ni 2 , Yuanyuan Dan 3., Sen Xu 4 1. school of Computer Science and Engineering, Jiangsu University of Science and Technology, Zhenjiang 212003. china 2. Department of Radiology, Carver College of medicine, The University of Iowa, Iowa City, IA 52242, USA 3. school of Biology and Chemical Engineering, Jiangsu University of Science and Technology, Zhenjiang 212003. china 4. school of Information Engineering, Yancheng Institute of Technology, Yancheng 224051, china school of Computer Science and Engineering Jiangsu University of Science and Technology Zhenjiang 212003 China Department of Radiology Carver College of Medicine The University of Iowa Iowa City IA 52242 USA school of Biology and Chemical Engineering Jiangsu University of Science and Technology Zhenjiang 212003 China school of Information Engineering Yancheng Institute of Technology Yancheng 224051 China

There have been many skewed cancer gene expression datasets in the post-genomic era. Extraction of differential expression genes or construction of decision rules using these skewed datasets by traditional algorithms will seriously underestimate the performance of the minority class, leading to inaccurate diagnosis in clinical trails. This paper presents a skewed gene selection algorithm that introduces a weighted metric into the gene selection procedure. The extracted genes are paired as decision rules to distinguish both classes, with these decision rules then integrated into an ensemble learning framework by majority voting to recognize test examples; thus avoiding tedious data normalization and classifier construction. The mining and integrating of a few reliable decision rules gave higher or at least comparable classification performance than many traditional class imbalance learning algorithms on four benchmark imbalanced cancer gene expression datasets.

关键词： cancer gene expression data class imbalance paired differential expression genes decision ruleensemble learning majority voting

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A PCA-based method for ancestral informative markers selection in structured populations

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Science china(Life Sciences) 2009年第10期52卷 972-976页

作者： ZHANG Feng1,3. ZHANG Lei1,3.& DENG Hong-Wen1,2,3.1 Key Laboratory of Biomedical Information Engineering of Ministry of Education, Institute of Molecular Genetics, school of Life Science and Technology, Xi’an Jiaotong University, Xi’an 710049, china 2 Institute of Bioscience and Biotechnology, school of Science, Beijing Jiaotong University, Beijing 100044, china 3.Departments of Orthopedic Surgery and basic Medical Science, school of medicine, University of Missouri-Kansas City, Kansas City, MO 64108, USA Key Laboratory of Biomedical Information Engineering of Ministry of Education Institute of Molecular Genetics School of Life Science and Technology Xi'an Jiaotong University Xi'an 710049 China.

Identification of population structure can help trace population histories and identify disease genes. Structured association (SA) is a commonly used approach for population structure identification and association mapping. A major issue with SA is that its performance greatly depends on the informa-tiveness and the numbers of ancestral informative markers (AIMs). Present major AIM selection meth-ods mostly require prior individual ancestry information, which is usually not available or uncertain in practice. To address this potential weakness, we herein develop a novel approach for AIM selection based on principle component analysis (PCA), which does not require prior ancestry information of study subjects. Our simulation and real genetic data analysis results suggest that, with equivalent AIMs, PCA-based selected AIMs can significantly increase the accuracy of inferred individual ancestries compared with traditionally randomly selected AIMs. Our method can easily be applied to whole genome data to select a set of highly informative AIMs in population structure, which can then be used to identify potential population structure and correct possible statistical biases caused by population stratification.

关键词： population structure principle component analysis ancestral informative markers

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A tentative study for the prediction of the CME related geomagnetic storm intensity and its transit time

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Science china(Technological Sciences) 2005年第6期48卷 648-668页

作者： ZHAO Xinhua1,2 & FENG Xueshang1,3.1. SIGMA Weather Group, Key Laboratory of Space Weather, Center for Space Science and Applied Re-search, Chinese Academy of Sciences, Beijing 100080, china 2. Graduate school of the Chinese Academy of Sciences, Beijing 10003., china 3. school of Geophysics and Geoinformation Systems, china University of Geosciences, Beijing 100083. china SIGMA Weather Group Key Laboratory of Space Weather Center for Space Science and Applied Research Chinese Academy of Sciences Beijing China Graduate school of the Chinese Academy of Sciences Beijing China school of Geophysics and Geoinformation Systems China University of Geosciences Beijing China

Using 80 CME-ICME events during 1997.1―2002.9, based on the eruptive source locations of CMEs and solar magnetic field observation at the photosphere, a current sheet magnetic coordinate (CMC) system is established in order to study the propagation of CME and its geoeffectiveness. In context of this coordinate system, the effect of the eruptive source location and the form of heliospheric current sheet (HCS) at the eruptive time of CME on the geomagnetic storm intensity caused by CME and the CME’s transit time at the Earth is investigated in detail. Our preliminary conclusions are: 1) The geomagnetic disturbances caused by CMEs tend to have the so-called “same side-opposite side effect”, i.e. CMEs erupt from the same side of the HCS as the earth would be more likely to arrive at the earth and the geomagnetic disturbances associated with them tend to be of larger magnitude, while CMEs erupting from the opposite side would arrive at the earth with less probability and the corresponding geomagnetic disturbance magnitudes would be relatively weaker. 2) The angular separation between the earth and the HCS affect the corresponding disturbance intensity. That is, when our earth is located near the HCS, adverse space weather events occur most probably. 3. The erupting location of the CME and its nearby form of HCS will also affect its arrival time at the earth. According to these conclusions, in this context of CMC coordinate we arrive at new prediction method for estimating the geomagnetic storm intensity (Dstmin) caused by CMEs and their transit times. The application of the empirical model for 80 CME-ICME events shows that the relative error of Dst is within 3.% for 59% events with Dstmin≤?50 nT, while the averaged absolute error of transit time is lower than 10 h for all events.

关键词： coronal mass ejections, current sheet magnetic coordinate system, geomagnetic storm intensity, transit time, prediction method.

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Cross-linked Polyethylenimine as Potential DNA Vector for Gene Delivery with High Efficiency and Low Cytotoxicity

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Acta Biochimica et Biophysica Sinica 2006年第11期38卷 780-787页

作者： Wei DONG~(1,2) Guang-Hui JIN~1 Shu-Feng LI~(1,3. Qi-Ming SUN~1 Ding-Yuan MA~1 Zi-Chun HUA~1 1 State Key Laboratory of Pharmaceutical Biotechnology,College of Life Science,Nanjing University,Nanjing 210093.china 2 school of Chemical Engineering Nanjing University of Science and Technology,Nanjing 210094,china 3.College of {3. Medical Science,Southeast University,Nanjing 210009,china State Key Laboratory of Pharmaceutical Biotechnology College of Life Science Nanjing UniversityNanjing 210093 China school of Chemical Engineering Nanjing University of Science and TechnologyNanjing 210094 China College of basic Medical Science Southeast UniversityNanjing 210009 China

Polyethylenimine (PEI) has been known as an efficient gene carrier with the highest cationiccharge *** transfection efficiency of PEI,along with its cytotoxicity,strongly depends on itsmolecular *** enhance its gene delivery efficiency and minimize cytotoxicity,we have synthesizedsmall cross-linked PEI with biodegradable linkages and evaluated their transfection efficiencies in *** study,branched PEI with a molecular weight of 800 Da was cross-linked by small diacrylate[1,4-butanediol diacrylate or ethyleneglycol dimethacrylate (EGDMA)] for 2-6 *** efficiencies of thecross-linked PEI in in vitro transfection of plasmid DNA containing enhanced green fluorescent protein(EGFP) reporter gene were assessed in melanoma B 16F10 cell line and other cell *** cytometrywas used to quantify the cellular entry efficiency of plasmid and the transgene expression *** of the cross-linked PEI in these cells were evaluated by MTT ***-PEI 800-4h,atypical cross-linked PEI reported here,mediated a more efficient expression of reporter gene than thecommercially available 25-kDa branched PEI control,and resulted in a 9-fold increase in gene deliveryin B16F10 cells and a 16-fold increase in 293. cells,while no cytotoxicity was found at the optimizedcondition for gene ***,the transfection activity of polyplexes was preserved in thepresence of serum proteins.

关键词： polyethylenimine gene delivery cytotoxicity B16F10 293. cell line

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