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The urban river embankment discuss the ecological constructionAnonymous XXXXXXXXAbstract: the urban river embankment construction as the object, discuss the current social background, analyses and compares the river embankment design of traditional methods and characteristics of ecological methods, and puts forward three modes of ecological design and their advantages and disadvantages, and expounds the present situation of the ecological construction in domestic bank and future : the bank; Ecology; Design way; Domestic situationText:A, backgroundRiver Banks part is the amphibious interlaced transition belt, has the remarkable edge effect. Here are active substances, nutrient and energy flow, offer a habitat for a variety of creatures. Natural state Banks often species richness, productivity traditional embankment design often single ?一、背景河流的堤岸部分是水陆交错的过渡地带，具有显著的边缘效应。这里有活跃的物质、养分和能量的流动，为多种生物提供了栖息地。自然状态下的堤岸往往物种丰富、生产力高。传统的堤岸设计往往会单纯从防洪角度出发，采用土堤或者土石混合堆砌起来高高的堤岸。它的优点在于高度的可靠性，结构设计后加起防护堤岸抗流水冲刷能力显著增强。对于洪水暴发频繁、侵蚀严重的区段，这样的设计无可厚非，而对于一般河流堤岸的修建，这样的设计则显得缺乏环境的美化和绿化，同时也破坏许多对生态起重要作用的自然因素，如破坏植被与河床间的联系，造成冲刷侵蚀转移等。另外，河流作为城市风貌不可多得的珍惜资源，也是城市风貌的特色要素，它的景观塑造显得十分必要。同时，堤岸景观建设必然使滨河地区土地价值提升，滨水开发的高投资回报的特点更增强了对城市堤岸景观建设的需求。二、需求——堤岸的生态化建设河流堤岸作为城市中最邻近河流的区域，是城市与河流的衔接线，它的景观规划是提高城市生活品质的需要，也是丰富城市景观的需要。生态化建设，它的根本思路是运用自然本身抗干扰和自我修复的能力来处理人与自然的关系。生态设计方法不同于传统用人工的结构和形式来取代自然的方法，而是用自然的结构和形式来顺应自然的进程。将河岸与河道在生态上联系起来，也就实现了物质、养分、能量的交流：对于生物，它提供了合适的栖息地；植物根系可固着土壤，枝叶可截留雨水，过滤地表迳流，抵抗流水冲刷，从而起到保护堤岸、增加堤岸结构的稳定性、净化水质、涵养水源的作用，而且随着时间的推移，这些作用被不断加强。同时，生态化建设以自然的外貌出现，容易与环境取得协调，造价也较低，不需要长期的维护管理。三、河流堤岸生态化设计方式河流堤岸生态化设计，要遵守生态设计的原则，注重地方性、保护与节约自然资本、让自然做功、显露自然，主要体现在对地域气候环境、河流地质地貌、水文变化的适应，对河流生态环境的考虑，对堤岸地形的处理和对筑堤材料的选择和构造方式方面。1） 人工类：传统方法是采用块石或混凝土块砖等堆砌。可在此基础上加以改进以适应河流景观设计的需求。a) 块石或混凝土块砖干砌，不用砂浆。这样在砌块之间就留有空隙，为后期滨河植物的生长提供了空间。随着时间的推移，堤岸会逐渐呈现出自然的风貌。b) 堤岸采用台阶式分级，台阶面上的空间加以利用，种植植物。当然这两种改进方法对于河岸处现有植被仍存在一定的不良影响，人工痕迹也过于明显。2） 自然类：充分利用堤岸植被原型，可直接将适用于滨河地带生长的植被种植于堤岸上，利用植物的根、茎、叶来稳固堤岸，防止侵蚀、控制沉积的同时也为生物提供了栖息地。3） 人工自然相结合综合了以上两种方法的优点，具有人工结构的稳定性和自然的外貌，见效快、生态效益好，以下为常见的两种类型：a) 种植植物的堆石将由大小不同的石块组成的堆石置于与水接触的土壤表面，再把活体切枝插入石堆中使斜坡更加稳定。根系可提高强度，植被可遮盖石块，使堤岸外貌更加自然。b) 与植物结合使用的插孔式混凝土块将预制的混凝土块以连锁的形式置于岸底的浅渠中，再将植物切枝或植株扦插于混凝土块之间和堤岸上部，其上覆土压实，再播种草本植物。堤岸生态化建设也存在一定的局限性。如：选用的材料及建造方法不同，堤岸的防护能力相差很大，需要运用多学科知识认真分析，这就为设计人员提出了更大的挑战；建造初期若受到强烈干扰，则会影响到以后防护作用的发挥等。这也就对河流堤岸的生态化设计提出了更高的要求。四、国内现状1）省会城市在我国省会城市及计划单列市中有近80％进行了堤岸景观规划。（参考文献[3]）城 市 项目名称 城 市 项目名称北 京 长河城市水系统综合治理 南 宁 堤岸园工程长 沙 湘江风光带 宁 波 滨江大道沿江景观工程成 都 府南河绿化工程 上 海 外滩、陆家嘴滨江大道福 州 闵江江滨公园 沈 阳 浑河观光旅游带广 州 珠江二沙段堤岸景观、芳村长堤建设 太 原 汾河公园贵 阳 南明河景观绿化工程 天 津 海河堤岸改造工程哈尔滨 松花江南岸沿江风景长廊 武 汉 汉口江滩一二期工程昆 明 盘龙江中段滨水生态景观建设 西 安 灞河大水大绿工程兰 州 黄河风情线 重 庆 南滨路滨江旅游观光大道从规划后建成情况看，这些城市河流堤岸景观项目都得到了当地政府与市民的肯定。在这些项目中，堤岸既可成为当地最具吸引力的城市公园，如太原的汾河公园和福州的江滨公园；堤岸也可成为市民日常休闲活动的热点地段，如南宁的堤路园和武汉的汉口江滩工程；堤岸还可成为城市最具特色的地段，如重庆的南滨路滨江旅游观光大道；堤岸更可成为城市旅游的热点，如上海的外滩和陆家嘴滨江大道。总之，经过景观规划的堤岸已成为当地最具特色的地区。从建设效果看，相对堤岸的原来面貌而言，统计资料中的这些景观工程都是较成功的，都成为当地城市关注的热点，成为当地政府的政绩工程，成为当地的民心工程。城市河流堤岸通过景观规划，有效地改善了滨河地段的环境，并带动滨河地段的开发。但必须清醒地认识到，这些城市堤岸景观项目规划并非尽善尽美，也存在这样或那样的问题，仍有待完善。2）中小城市城市经济实力的强大决定了其城市建设水平的高标准和高水平。中小城市河流堤岸景观与统计资料中的城市存在较大的差距，存在更多的问题。特别是由于资金问题，堤岸景观是，纯人工，状态的钢筋混凝土防洪堤，或保持自然防洪状态的土石堤，没有经过景观规划，易造成城市资源的极大浪费。五、前景目前，河流景观建设，特别是城市河流景观建设，在中国正方兴未艾；在发达国家中也是一个久盛不衰的话题。 回顾发达国家河流景观建设的历史，自20世纪70年代以来，随着人们环境意识的普遍增强，重视河流景观的生态功能已成为一个时代的呼唤，河流景观建设的生态设计方法也已得到了空前的重视和发展。他山之石可以攻玉，借鉴发达国家已经形成的成熟的理念和做法，可以使我们少走弯路，搭上隆隆前进的生态建设之车。
Summary of the study on construction project management methodology: is the largest number of construction projects, the most typical "projects", construction project management practice is one of the important sources of project management theory with integrated project management system of project management theory and practical experience provide theories for construction project management tools, making increasingly systematic and scientific management of construction projects. Tags: 1, raised issues of project management and overview of project management is becoming a modern social address "one-off questions" effective tools that a kind of professional qualification, wide application in the process of social, economic and rapid development is that it provides a new way. Modern social economic total constantly increased, economic globalization, and information trend increasingly enhanced, development speed speed up, process complex, new of industry, and field constantly appears, products development cycle shortened, led increasingly more of "one-time", and no precedent can through of task of appears, especially enterprise and management social public affairs of Government, more faced was increasingly complex of from market and at home and abroad Affairs of various problem, involves of factors, and interests main increasingly more, these problem General of management approach difficult to solution or successfully solution, and from construction project and the military, and space exploration, field of project management method by upgrade for project management theory and thought, provides has solution social economic development non-General problem of means. You can think, lies in its development and application of project management method. Widely promoted popular project management in China in recent years, the ascendant, and we introduce, universal focus, should not be a complicated theories, concepts, procedures, and should be concise, applicable and effective way. Regardless of the economic structure changes, construction remains the main carrier of China's economic development. In comprehensively building a comfortably well-off society and development in China under the background of the Northwest, Northeast, become important means for economic revitalization in many parts of the project. Some cities even proposed "project will flourish cities" slogan. Rely mainly on the application of new scientific and technological achievements in economic progress, relying on the content-type extension, depend on the situation of knowledge innovation, rely on an extensive project is not a suitable means of revitalizing the economy, has been in China for failure to pay the investment decision of enormous cost, so using the scientific method on engineering construction project management, including assessment of argumentation, is the important condition of the building can achieve the desired purpose of the project. Project management methods are not only specific methods, including thought. Party of 16 session third plenary made people, and can continued of science development view, "people", and "can continued" of a basic content is science and technology, and economic, and environment, and social of coordination development, project construction do for development economic of important means, not only cost huge, and on around environment has larger effect, so must meet can continued, and coordination development requirements, this is project management of important guiding ideology one. Methodology is the study of methods. Comprehensively building a comfortably well-off society and development in Northwest and northeast of real background might set off a new round of upsurge of construction, sustainable and coordinated development of scientific concept of development put certain constraints on the construction of macro-control, this method should be implemented in construction project management. Project management is not only for the purposes of achieving specific objectives (duration, quality and cost), should also do project construction and coordination of environmental, social or risk repeating previous errors, mistake, got out of the investment mistakes. This is also a project management method to rise to "methodology" meaning one. Project management of theory so far has can said is is complete of, this not only performance in rich diverse of theory results as various project management books (including translation) of launched, more performance in this area has has own of "knowledge system", as United States Project Management Association (PMI) Yu 1987 made and by 1996, and 2000 amendment of "project management knowledge system", international project management Association 1997 launched has "project management personnel ability benchmark", China double method society for the project management Research Committee also Yu 2001 launched China project management knowledge system under. Each of these knowledge systems project management body of knowledge is divided into a number of areas, elements, modules, China knowledge system "methods and tools" concept, other systems do not have on the topic independent of the method, but throughout all stages of project management, processes, fields. Various project management works also focus on content, stage of project management, program, organization of expositions devoted to rare. Of course, this does not mean to ignore, perfection but summary, concentrate on methods of research and contribute to the popularization and application of project management project management role into full play.建设工程项目管理的方法论研究摘要：建设工程项目是数量最多、最典型的“项目”，建设工程项目管理的实践是项目管理理论的重要渊源之一，融合了项目管理实践经验的系统的项目管理理论又为建设工程项目管理提供了理论工具，使得建设工程项目管理日益系统化、科学化。 关键词：项目管理 工程1、问题的提出及现状综述 项目管理之所以成为一种现代社会解决“一次性问题”的有效工具以至于一种职业资格，在社会事务、经济过程中广泛应用并且迅速发展，就在于它提供了一种新的方法。现代社会经济总量不断增加，经济全球化、信息化趋势日益增强，发展速度加快，过程复杂，新的行业、领域不断出现，产品开发周期缩短，导致越来越多的“一次性”、无先例可循的任务的出现，尤其是企业和管理社会公共事务的政府，更面临着日趋复杂的来自市场和国内外事务的各种问题，涉及到的因素、利益主体越来越多，这些问题常规的管理办法难以解决或圆满解决，而来自建设工程项目及军事、空间探索等领域的项目管理方法经提升为项目管理理论和思想，提供了解决社会经济发展中非常规问题的手段。可以认为，项目管理的发展与应用在于它的方法。项目管理在中国广泛推广普及是近几年的事，方兴未艾，而我们引进、普及的重点，不应是繁琐的理论、概念、程序，而应该是简明、适用、有效的方法。 不论经济结构如何变化，工程项目建设仍是我国经济发展的主要载体。在我国目前全面建设小康社会、开发西北、振兴东北的背景下，项目建设成为许多地区经济振兴的重要手段。有的城市甚至提出“项目兴市”的口号。在经济进步主要依靠新科技成果应用、依靠内涵式扩展、依靠知识创新的形势下，靠粗放式项目建设并不是合适的振兴经济的手段，我国也一直在为投资决策的失误付出巨大代价，所以利用科学方法对工程建设项目进行管理，包括评估论证，将是项目建设能否达到预期目的的重要条件。 工程项目管理方法不仅是具体方法，也包括思想。党的十六届三中全会提出以人为本、可持续的科学发展观，“以人为本”、“可持续”的一个基本内容就是科学技术、经济、环境、社会的协调发展，工程项目建设做为发展经济的重要手段，不仅耗资巨大，而且对周围环境有较大影响，所以必须满足可持续、协调发展要求，这也是工程项目管理的重要指导思想之一。方法论是对方法的研究。全面建设小康社会、开发西北、振兴东北的现实背景可能掀起新一轮工程建设热潮，可持续、协调发展科学发展观对工程项目建设提出了一定制约的宏观控制，这一点要落实到建设项目管理的方法上。工程项目管理的目的不仅是实现具体的目标（工期、质量、费用），也应做到项目建设与环境、社会的协调，否则可能重蹈以前投资失误、失策、失控的覆辙。这也是工程项目管理方法上升到“方法论”的意义之一。 项目管理的理论至今已可以说是很完备的，这不仅表现在丰富多样的理论成果如各种项目管理书籍（包括翻译）的面世，更表现在这个领域有了自己的“知识体系”，如美国项目管理协会（PMI）于1987年提出并经1996、2000年修订的“项目管理知识体系”，国际项目管理协会1997年推出了“项目管理人员能力基准”，中国双法研究会项目管理研究委员会也于2001年推出《中国项目管理知识体系》。上述各知识体系将项目管理知识划分为若干领域、要素、模块，中国的知识体系有“方法与工具”概念，其他体系未有关于方法的独立专题，而是贯穿于项目管理的各个阶段、过程、领域。各种项目管理著作也多侧重于对项目管理内容、阶段、程序、组织等的论述，对于方法的专门论述不多见。当然这并不意味着对方法的忽略，但对方法的总结、集中研究与完善有助于项目管理的推广应用的项目管理作用的充分发挥。
土木工程专业的英文论文格式
导语：土木工程专业的英文的论文格式包括哪些内容呢?土木工程是建造各类工程设施的科学技术的统称。下面是我分享的土木工程专业的英文的论文格式，欢迎阅读!
土木工程专业的英文论文格式均以美国土木工程师协会出版社发布的标准格式为准。
英语论文用激光打印机打印，打印稿为黑白稿，彩色打印件会影响出版效果。
版心：a4纸，上、下页边距 cm，左、右页边距均为 mm。论文内容宽不得超过， 长不得超过。
字体和字号：正文，标题，作者联络信息和图表中的文字均为times new roman 12号字。可以跟据需要使用同类字体中的粗体，斜体。
行距：单倍行距。
页码： 论文正文和文后所附图例都需添加页码。页码为阿拉伯数字，位于页面下方居中。
文体： 文章应语法正确，技术用词准确。标题应该以最简洁的语言概括文章内容。如果标题较长，请采用title: subtitle的形式。
数学公式：文中的数学公式不得手写，必须打印。公式如果在文中多次被引用，应该编号。公式之间，公式和正文之间都应该空一行。 单位： 文中所用的度量衡单位应为国际单位。可在括号内，单位对应表中列出其他单位。有关国际单位的使用(standard practice for use of the international system of units)可以通过电话1-向asce索取。其他相关使用参考文献，如anmc metric editorial guide， 5th ed，1992 可向美国国家公制协会 索取(american national metric council， 1735n. lynn street， suite 950， arlington， va 22209-2022)
图表：
标题说明和图例：插入的图表应该以出现顺序编号(figure 1，figure 2，table 1，table 2)。图的说明和标题，包括图的序号应该位于图的下方。表的说明和标题，包括表的序号应该位于表格上方。
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参考文献： 所有参考文献为单倍行距，放在文章最后，按照第一作者姓氏的字母顺序排列。如有同一作者的两篇以上文献，按出版年代先后排列。正文中引用参考文献时，作者和出版年代应该放入括号内。由于上标缩印后会变小，难于辨认，正文中不使用上标标注参考文献。所列出的参考文献应当在正文中都有所引用，如果正文中没有引用，请将文献列入文章最后的附加信息(additional information)部分，或者相关材料(related materials)部分。
1.《建筑结构制图标准》GB/T50105-2001
2.《建筑结构荷载规范》GB5009-2001(2006版)
3.《混凝土结构设计规范》GB50010-2002
4.《建筑地基基础设计规范》GB5007-2002
5.《砌体结构设计规范》GB5003-2001
6.《建筑抗震设计规范》GB5011-2001
7.《钢结构设计规范》GB50017-2003
8.《建筑结构构造资料》(合订本)，中国建筑工业出版社，1998年。
9.《混凝土结构构造手册》，中国建筑工业出版社，2002年。
10.《地基基础设计手册》，上海科技出版社，1998年。
11.《混凝土结构设计手册》，中国建筑工业出版社，2002年。
12.《建筑结构静力计算手册》，中国建筑工业出版社，1999年
13.《建筑结构强制性国家标准》(简装本)，中国建筑工业出版社，2001年9月
14. 任全宏、常建军.钢筋混凝土多层框架结构房屋结构设计中应注意的几个问题。陕西建筑2007，145(7).
15. 范俊梅. 钢筋混凝土多层框架结构设计问题分析. 科技资讯2008，3.
16. Basic Principles for Reinforced Concrete Structure Design
17.建筑、结构设计有关图集资料以及专业课程教材等。
1、《工业建筑》
《工业建筑》是中国钢铁工业协会主管，中冶建筑研究总院有限公司主办的综合性建筑科学类科技期刊；报道内容涵盖土木建筑领域的主要学科，包括建筑学、建筑结构、岩土工程、建筑材料、建筑施工五大专业。据2018年5月《工业建筑》编辑部官网显示，《工业建筑》编辑委员会拥有顾问17人，编委65人。
2、《建筑经济》
《建筑经济》是中华人民共和国住房和城乡建设部主管，中国建筑学会、中国建筑设计研究院、亚太建设科技信息研究院主办的建筑经济与管理领域主流理论期刊。《建筑经济》创刊于1980年，为季刊；中国知网显示，1982年改为双月刊，从1985年开始改为月刊。 据2018年5月《建筑经济》官网显示，《建筑经济》编委会拥有43位编委。
3、《中国农村水利水电》
《中国农村水利水电杂志》（China Rural Water and Hydropower）是由中华人民共和国水利部主管，水利部中国灌溉排水发展中心、水利部农村水电及电气化发展局、武汉大学、 中国国家灌溉排水委员会主办的学术性期刊。《中国农村水利水电杂志》创刊于1959年。据2018年5月《中国农村水利水电杂志》官网显示，《中国农村水利水电杂志》编委会拥有编委29人。
4、《建筑科学》
《建筑科学》1985年创刊，是由中华人民共和国住房和城乡建设部主管、中国建筑科学研究院有限公司主办的建筑科学类综合性技术期刊。据2020年第06期期刊内页显示，《建筑科学》编委会拥有编辑1人、责任编辑1人、美编1人，主编1人、副主编1人、执行主编1人。
5、《城市发展研究》
《城市发展研究》创刊于1994年，是由中国科学技术协会和中华人民共和国建设部共同主管、中国城市科学研究会主办的经济与管理科学期刊。截至2011年7月《城市发展研究》编辑部官网显示，《城市发展研究》编辑委员会顾问编委有8人、编委委员有44人。
好可惜，我不会翻译。
《工业工程》官方描述：“It publishes original contributions on the development of new computerized methodologies for solving industrial engineering problems, as well as the applications of those methodologies to problems of interest in the broad industrial engineering and associated communities. The journal encourages submissions that expand the frontiers of the fundamental theories and concepts underlying industrial engineering techniques.”“本期刊发布关于开发新的计算机化方法以解决工业工程问题的原始贡献，以及这些方法在广泛的工业工程和相关社区感兴趣的问题的应用。 本期刊鼓励扩大工业工程技术基础理论和前沿概念的提交。”偏重的研究方向： 管理科学与工程(11) 管理综合(11) 工业工程与管理(6) 工业工程(3) 信息科学(3) 运筹(2) 工程与材料(2) 自动化(2) 运筹与管理(2) 系统科学与系统工程(2) 算法(2) industrial engineering(1) 计算机仿真(1) 水利科学与海洋工程(1) 水文、水资源(1) 风险管理技术与方法(1) 数理科学(1) 数学(1) 运筹学(1) 机械工程(1) 计算机科学(1) 机械动力学(1) 信息系统与管理(1) 计算机应用技术(1)
权威的可能了解的不多，只有一本（土木工程）有些了解的
已发，请查收，希望能帮到你，至于翻译得靠你自己了，急用的话多加点分悬赏，这样才有更多的知友及时帮助你
Production Line Balancing——Gavriel SalvendyThe scope of this study is to explore the understanding of Production-line Manufacturing and Balancing, Types of Line Balancing, Equipment Balancing and its Failure and Analysis. A production line is said to be in balance when every worker’s task takes the same amount of time. Line balancing is a manufacturing-engineering function in which whole collection of production-line tasks are divided into equal portions. Well-balanced lines avoid labour idealness and improve productivity. Production Line Balancing Line-balancing strategy is to make production lines flexible enough to absorb external and internal irregularities. There are two types of line balancing, which we have explained as： Static Balance – Refers to long-term differences in capacity over a period of several hours or longer. Static imbalance results in underutilization of workstations, machines and people. Dynamic Balance – Refers to short-term differences in capacity, like, over a period of minutes, hours at most. Dynamic imbalance arises from product mix changes and variations in work time unrelated to product mix. Labour Balancing and AssignmentsStrategy of production line stability is the tendency for labour assignments to be fixed. Labour feasibility is an important feature in the strategy of production line flexibility linked to individual skills and capabilitiesWhen one worker is having problem in performing his assigned task and experiencing delay due to technical problem(s), other worker(s) should move into help. The management practice of deliberately pulling worker’s of the line when the line is running smoothly. The movement of whole crews from one dedicated line to another as the model mix changes. Group Technology – In which one worker can handle variety of tasks (automation) in a single work centre. Equipment Balancing While balancing equipment, attempt to ensure that each piece of equipment in the work cell has the same amount of work. Now days every manufacturer is attempting to maximize the utilization of all available equipments. Such high utilization is often counterproductive and may be the wrong goal because; high utilization is usually accompanied by high inventory. Equipment Failure An equipment failure is a major serious matter, with the potential to shut down a production line. To avoid such failures one should not overload the equipments, and workers should be trained to perform a daily machine checking (preventive maintenance) and following standard operating procedures. The advantage for Maintenance and Engineering Department does not lie in running late shifts, hence calculate the preventive maintenance time and schedule the activity. Analysis Analysis is generally performed by Competent Technical Staff. Begin the analysis with division of production-line work into small tasks, determination of task time standards, specification of required task sequencing and notation of constraints. If bottle neck task is in the way of good balance, the Competent Technical Staff should analyze the task to reduce the time it takes to Balancing LeadershipWorkmen should lead the production line balancing effort, so that they can react quickly when line imbalances (static and dynamic) crop up as a result of changeover to make a different item or changes in the output Production-line balancing study tends to employ thought and ingenuity to change conditions. Production-line design and operation is more art than science. Labour flexibility is the key to effective resource management. The idea of worker’s checking and doing minor repair work on their own equipment possibly decreases the risk of equipment failure. Selecting an appropriate set of balancing mechanism is a part of work cell design and it must be linked with many other decisions for the system to function well. Keywords: line balancing industrial equipment failure
我有一份《The SMED System》，你要不要？讲的是如何快速换模（换线或叫转机、转拉）
把这个给我发一份行吗，谢谢!
The urban river embankment discuss the ecological constructionAnonymous XXXXXXXXAbstract: the urban river embankment construction as the object, discuss the current social background, analyses and compares the river embankment design of traditional methods and characteristics of ecological methods, and puts forward three modes of ecological design and their advantages and disadvantages, and expounds the present situation of the ecological construction in domestic bank and future : the bank; Ecology; Design way; Domestic situationText:A, backgroundRiver Banks part is the amphibious interlaced transition belt, has the remarkable edge effect. Here are active substances, nutrient and energy flow, offer a habitat for a variety of creatures. Natural state Banks often species richness, productivity traditional embankment design often single ?一、背景河流的堤岸部分是水陆交错的过渡地带，具有显著的边缘效应。这里有活跃的物质、养分和能量的流动，为多种生物提供了栖息地。自然状态下的堤岸往往物种丰富、生产力高。传统的堤岸设计往往会单纯从防洪角度出发，采用土堤或者土石混合堆砌起来高高的堤岸。它的优点在于高度的可靠性，结构设计后加起防护堤岸抗流水冲刷能力显著增强。对于洪水暴发频繁、侵蚀严重的区段，这样的设计无可厚非，而对于一般河流堤岸的修建，这样的设计则显得缺乏环境的美化和绿化，同时也破坏许多对生态起重要作用的自然因素，如破坏植被与河床间的联系，造成冲刷侵蚀转移等。另外，河流作为城市风貌不可多得的珍惜资源，也是城市风貌的特色要素，它的景观塑造显得十分必要。同时，堤岸景观建设必然使滨河地区土地价值提升，滨水开发的高投资回报的特点更增强了对城市堤岸景观建设的需求。二、需求——堤岸的生态化建设河流堤岸作为城市中最邻近河流的区域，是城市与河流的衔接线，它的景观规划是提高城市生活品质的需要，也是丰富城市景观的需要。生态化建设，它的根本思路是运用自然本身抗干扰和自我修复的能力来处理人与自然的关系。生态设计方法不同于传统用人工的结构和形式来取代自然的方法，而是用自然的结构和形式来顺应自然的进程。将河岸与河道在生态上联系起来，也就实现了物质、养分、能量的交流：对于生物，它提供了合适的栖息地；植物根系可固着土壤，枝叶可截留雨水，过滤地表迳流，抵抗流水冲刷，从而起到保护堤岸、增加堤岸结构的稳定性、净化水质、涵养水源的作用，而且随着时间的推移，这些作用被不断加强。同时，生态化建设以自然的外貌出现，容易与环境取得协调，造价也较低，不需要长期的维护管理。三、河流堤岸生态化设计方式河流堤岸生态化设计，要遵守生态设计的原则，注重地方性、保护与节约自然资本、让自然做功、显露自然，主要体现在对地域气候环境、河流地质地貌、水文变化的适应，对河流生态环境的考虑，对堤岸地形的处理和对筑堤材料的选择和构造方式方面。1） 人工类：传统方法是采用块石或混凝土块砖等堆砌。可在此基础上加以改进以适应河流景观设计的需求。a) 块石或混凝土块砖干砌，不用砂浆。这样在砌块之间就留有空隙，为后期滨河植物的生长提供了空间。随着时间的推移，堤岸会逐渐呈现出自然的风貌。b) 堤岸采用台阶式分级，台阶面上的空间加以利用，种植植物。当然这两种改进方法对于河岸处现有植被仍存在一定的不良影响，人工痕迹也过于明显。2） 自然类：充分利用堤岸植被原型，可直接将适用于滨河地带生长的植被种植于堤岸上，利用植物的根、茎、叶来稳固堤岸，防止侵蚀、控制沉积的同时也为生物提供了栖息地。3） 人工自然相结合综合了以上两种方法的优点，具有人工结构的稳定性和自然的外貌，见效快、生态效益好，以下为常见的两种类型：a) 种植植物的堆石将由大小不同的石块组成的堆石置于与水接触的土壤表面，再把活体切枝插入石堆中使斜坡更加稳定。根系可提高强度，植被可遮盖石块，使堤岸外貌更加自然。b) 与植物结合使用的插孔式混凝土块将预制的混凝土块以连锁的形式置于岸底的浅渠中，再将植物切枝或植株扦插于混凝土块之间和堤岸上部，其上覆土压实，再播种草本植物。堤岸生态化建设也存在一定的局限性。如：选用的材料及建造方法不同，堤岸的防护能力相差很大，需要运用多学科知识认真分析，这就为设计人员提出了更大的挑战；建造初期若受到强烈干扰，则会影响到以后防护作用的发挥等。这也就对河流堤岸的生态化设计提出了更高的要求。四、国内现状1）省会城市在我国省会城市及计划单列市中有近80％进行了堤岸景观规划。（参考文献[3]）城 市 项目名称 城 市 项目名称北 京 长河城市水系统综合治理 南 宁 堤岸园工程长 沙 湘江风光带 宁 波 滨江大道沿江景观工程成 都 府南河绿化工程 上 海 外滩、陆家嘴滨江大道福 州 闵江江滨公园 沈 阳 浑河观光旅游带广 州 珠江二沙段堤岸景观、芳村长堤建设 太 原 汾河公园贵 阳 南明河景观绿化工程 天 津 海河堤岸改造工程哈尔滨 松花江南岸沿江风景长廊 武 汉 汉口江滩一二期工程昆 明 盘龙江中段滨水生态景观建设 西 安 灞河大水大绿工程兰 州 黄河风情线 重 庆 南滨路滨江旅游观光大道从规划后建成情况看，这些城市河流堤岸景观项目都得到了当地政府与市民的肯定。在这些项目中，堤岸既可成为当地最具吸引力的城市公园，如太原的汾河公园和福州的江滨公园；堤岸也可成为市民日常休闲活动的热点地段，如南宁的堤路园和武汉的汉口江滩工程；堤岸还可成为城市最具特色的地段，如重庆的南滨路滨江旅游观光大道；堤岸更可成为城市旅游的热点，如上海的外滩和陆家嘴滨江大道。总之，经过景观规划的堤岸已成为当地最具特色的地区。从建设效果看，相对堤岸的原来面貌而言，统计资料中的这些景观工程都是较成功的，都成为当地城市关注的热点，成为当地政府的政绩工程，成为当地的民心工程。城市河流堤岸通过景观规划，有效地改善了滨河地段的环境，并带动滨河地段的开发。但必须清醒地认识到，这些城市堤岸景观项目规划并非尽善尽美，也存在这样或那样的问题，仍有待完善。2）中小城市城市经济实力的强大决定了其城市建设水平的高标准和高水平。中小城市河流堤岸景观与统计资料中的城市存在较大的差距，存在更多的问题。特别是由于资金问题，堤岸景观是，纯人工，状态的钢筋混凝土防洪堤，或保持自然防洪状态的土石堤，没有经过景观规划，易造成城市资源的极大浪费。五、前景目前，河流景观建设，特别是城市河流景观建设，在中国正方兴未艾；在发达国家中也是一个久盛不衰的话题。 回顾发达国家河流景观建设的历史，自20世纪70年代以来，随着人们环境意识的普遍增强，重视河流景观的生态功能已成为一个时代的呼唤，河流景观建设的生态设计方法也已得到了空前的重视和发展。他山之石可以攻玉，借鉴发达国家已经形成的成熟的理念和做法，可以使我们少走弯路，搭上隆隆前进的生态建设之车。
你是哪个省的啊 看看如果可以的话 可以给你一份全面的
有一篇施工监控的论文，你查收一下吧，希望对你有用！
童鞋你好！这个估计需要自己搜索了！网上基本很难找到免费给你服务的！我在这里给你点搜索国际上常用的外文数据库：----------------------------------------------------------❶ISI web of knowledge Engineering Village2❷Elsevier SDOL数据库 IEEE/IEE(IEL)❸EBSCOhost RSC英国皇家化学学会❹ACM美国计算机学会 ASCE美国土木工程师学会❺Springer电子期刊 WorldSciNet电子期刊全文库❻Nature周刊 NetLibrary电子图书❼ProQuest学位论文全文数据库❽国道外文专题数据库 CALIS西文期刊目次数据库❾推荐使用ISI web of knowledge Engineering Village2-----------------------------------------------------------中文翻译得自己做了，实在不成就谷歌翻译。弄完之后，自己阅读几遍弄顺了就成啦！学校以及老师都不会看这个东西的！外文翻译不是论文的主要内容！所以，很容易过去的！祝你好运！
生物基因工程论文参考文献汇总 生物基因工程论文参考文献怎么写?有哪些格式要求，下面我就为大家推荐一些优秀的范例，希望大家喜欢![1] Brackett B G, Baranska W, Sawicki W，et al. Uptake of heterologous genome by mammalianspermatozoa and its transfer to ova through fertilization. Proc Natl Acad Sci USA,1971,68(2):353-357. [2] Jaenisch R, Mintz B. Simian virus 40 DNA sequences in DNA of healthy adult mice derived frompreimp antation blastocysts injected with viral DNA. Proc Natl Acad Sci USA, 1974,71 (4): 1250-1254. [3] Palmiter R D, Brinster R L, Hammer R E, et al. Dramatic growth of mice that develop from eggsmicroinjected with metallothionein-growth hormone fusion genes. Nature, 1982,300(5893):611-615. [4] 李宁.动物克隆与基因组编辑.中国农业大学出版社，2012. [5] Hammer R E, Pursel V G, Rexroad C J, et al. Production of transgenic rabbits, sheep and pigs bymicroinjection. Nature, 1985,315(6021):680-683 [6] 杜伟，崔海信，王 琰 ，等.精子载体法制备转基因动物的'研究进展.生物技术通报，2012(12):13-18. [7] Maione B，Lavitrano M, Spadafora C, et al. Sperm-mediated gene transfer in mice. Mol ReprodDev, 1998,50(4):406-409. [8] Lavitrano M, Bacci M L, Forni M, et al. Efficient production by sperm-mediated gene transfer ofhuman decay accelerating factor (hDAF) transgenic pigs for xenotransplantation. Proc Matl Acad SciUSA, 2002,99(22):14230-14235. [9] Sperandio S, Lulli V，Bacci M L, et al. Sperm - mediated DNA transfer in bovine and swinespecies. Animal biotechnology, 1996,7(1):59-77. [10] 武坚，刘明军，李文蓉，等.精子载体介导法生产转基因绵羊的研究.草食家畜，2001(S2):186-190. [11] Pfeifer A, Kessler T, Yang M, et al. Transduction of liver cells by lentiviral vectors: analysis inliving animals by fluorescence imaging. Mol Ther，2001,3(3):319-322. [12] Lois C, Hong E J, Pease S, et al. Germline transmission and tissue-specific expression oftransgenes delivered by lentiviral vectors. Science, 2002,295(5556):868-872. [13] Hofmann A, Kessler B, Ewerling S，et al. Efficient transgenesis in farm animals by lentiviralvectors. EMBO Rep, 2003,4( 11): 1054-1060. [14] Hofmann A, Zakhartchenko V, Weppert M, et al. Generation of transgenic cattle by lentiviral genetransfer into oocytes’ Biol Reprod, 2004,71 (2):405-409 [15] Lillico S G, Sherman A, McGrew M J，et al. Oviduct-specific expression of two therapeuticproteins in transgenic hens. Proc Natl Acad Sci USA，2007,104(6): 1771-1776. [16] Lyall J，Irvine R M, Sherman A, et al. Suppression of avian influenza transmission in geneticallymodified chickens. Science，2011,331(6014):223-226. [17] Golding M C, Long C R，Carmell M A, et al. Suppression of prion protein in livestock by RNAinterference. Proc Natl Acad Sci USA, 2006,103(14):5285-5290. [18] 杨春荣，窦忠英.利用干细胞生产转基因动物研究进展.西北农林科技大学学报(自然科学版)，2006(07):37-40. [19] Hai T, Teng F，Guo R, et al. One-step generation of knockout pigs by zygote injection ofCRISPR/Cas system. Cell Res, 2014,24(3):372-375. [20] Hongbing H, Yonghe M A, Tao W, et al. One-step generation of myostatin gene knockout sheepvia the CRISPR/Cas9 system. Frontiers of Agricultural Science and Engineering, 2014,1(1):2-5. [21] Swanson M E，Martin M J, O'Donnell J K, et al. Production of functional human hemoglobin intransgenic swine. Biotechnology (N Y)，1992,10(5):557-559. [22] Zbikowska H M，Soukhareva N, Behnam R, et al. Uromodulin promoter directs high-levelexpression of biologically active human alpha 1-antitrypsin into mouse urine. Biochem J, 2002,365(Pt1):7-11. [23] Golovan S P，Hayes M A, Phillips J P，et al. Transgenic mice expressing bacterial phytase as amodel for phosphorus pollution control. Nat Biotechnol, 2001,19(5):429-433. [24] Rapp J C, Harvey A J, Speksnijder G L, et al. Biologically active human interferon alpha-2bproduced in the egg white of transgenic hens. Transgenic Res, 2003,12(5):569-575. [25] Wright G, Carver A, Cottom D, et al. High level expression of active human alpha-1 -antitrypsin inthe milk of transgenic sheep. Biotechnology (N Y), 1991,9(9):830-834. [26] Li S, Ip D T, Lin H Q, et al. High-level expression of functional recombinant humanbutyrylcholinesterase in silkworm larvae by Bac-to-Bac system. Chem Biol Interact,2010,187(1-3):101-105. [27] 刘英，张瑞君，伍志伟，等.转基因疾病动物模型的研究进展.动物医学进展，2006(12):44-49. [28] Kragh P M, Nielsen A L, Li J, et al. Hemizygous minipigs produced by random gene insertion andhandmade cloning express the Alzheimer's disease-causing dominant mutation APPsw. Transgenic Res,2009,18(4):545-558. [29] Lee M K, Stirling W, Xu Y, et al. Human alpha-synuclein-harboring familial Parkinson'sdisease-linked Ala-53 Thr mutation causes neurodegenerative disease with alpha-synucleinaggregation in transgenic mice. Proc Natl Acad Sci USA, 2002,99(13):8968-8973. ;
summary：With the current molecular technology rapid development, the technology used in our life has more and more. For this kind of magical technology, people hei mixed, this debate. Especially in recently, American shows that by gene engineering technology has developed artificial blood and is scheduled to 2013 after artificial blood for human trials, but also aroused people's controversial, and this paper is mainly to the use of genetic engineering to create artificial blood to a shallow prohibited.
Synthesis of optically pure ethyl (S)-4-chloro-3-hydroxybutanoateby Escherichia coli transformant cells coexpressingthe carbonyl reductase and glucose dehydrogenase genes由共表达碳酰还原酶和葡萄糖脱氢酶的大肠杆菌转化细胞合成纯光学（S）-4-氯-3-羟基丁酸乙酯Abstract The asymmetric reduction of ethyl 4-chloro-3-oxobutanoate (COBE) to ethyl (S)-4-chloro-3-hydroxybutanoate((S)-CHBE) was investigated. Escherichia coli cells expressing both the carbonyl reductase (S1) gene from Candida magnoliae and the glucose dehydrogenase (GDH) gene from Bacillus megaterium were used as thecatalyst. In an organic-solvent-water two-phase system,(S)-CHBE formed in the organic phase amounted to M (430 g/l), the molar yield being 85%. E. coli transformant cells coproducing S1 and GDH accumulated M (208 g/l) (S)-CHBE in an aqueous monophase system by continuously feeding on COBE, which is unstable in an aqueous solution. In this case, the calculated turnover of NADP+ (the oxidized form of nicotinamide adenine dinucleotide phosphate) to CHBE was 21,600 mol/mol. The optical purity of the (S)-CHBE formed was 100% enantiomeric excess in both systems. The aqueous system used for the reduction reaction involving E. coli HB101 cells carrying a plasmid containing the S1 and GDH genes as a catalyst is simple. Furthermore, the system does not require the addition of commercially available GDH or an organic solvent. Therefore this system is highly advantageous for the practical synthesis of optically pure (S)-CHBE.本本篇文献研究了利用COBE不对称合成（S）-4-氯-3-羟基丁酸乙酯（CHBE）。大肠杆菌细胞作为催化剂同时表达了来自念珠菌属magnoliae的碳酰还原酶和来自巨大芽孢杆菌的葡萄糖脱氢酶基因。在水/有机溶剂两相体系中，(S)-CHBE在有机相中的浓度可以达到（430g/l），摩尔产率达到85%。大肠杆菌的副产物S1和GDH也达到了（208g/l），COBE在水相中不稳定，所以(S)-CHBE可以在水单相中不停的生成。在这种情况下，适当的从NADP+到CHBE的转变达到了21,600 mol/mol。所形成的CHBE的旋光度在这种体系中100%对映体过量。在水相中用携带含有S1和GDH基因质粒的E. coli HB101作为催化剂不对称还原是比较简单的。并且，这种体系并不额外需要商业GDH或者有机溶剂。因此，这种体系对于实际合成纯光学活性的(S)-CHBE是非常方便的。Optically active 4-chloro-3-hydroxybutanoic acid esters are useful chiral building blocks for the synthesis of pharmaceuticals. The (R)-enantiomer is a precursor of L-carnitine (Zhou et al. 1983), and (S)-enantiomer is an important starting material for hydroxymethylglutaryl- CoA (HMG-CoA) reductase inhibitors (Karanewsky et al. 1990). Many studies have described the microbial or enzymatic asymmetric reduction of 4-chloro-3-oxobutanoic acid esters (Aragozzini and Valenti 1992; Bare et ; Hallinan et al. 1995; Patel et al. 1992; Shimizu et al. 1990; Wong et al. 1985) based on the reduction by baker’s yeast (Zhou et al. 1983).We have previously showed that Candida magnoliae AKU4643 cells reduced ethyl 4-chloro-3-oxobutanoate (COBE) to (S)-CHBE with an optical purity of 96% enantiomeric excess (.) (Yasohara et al. 1999). As this yeast has at least three different stereoselective reductases (Wada et al. 1998, 1999a, b), the (S)-CHBE produced by this yeast was not optically pure. From among these three enzymes, an NADPH-dependent carbonyl reductase, designated as S1, was purified and characterized in some detail (Wada et al. 1998). We cloned and sequenced the gene encoding S1 and overexpressed it in Escherichia coli cells. This E. coli transformant reduced COBE to optically pure (S)-CHBE in the presence of glucose, NADP+, and commercially available glucose dehydrogenase (GDH) as a cofactor generator (Yasoharaet al. 2000). Here, we describe the construction of three E. coli transformants coexpressing the S1 from C. magnoliae and GDH from Bacillus megaterium genes and analyze the reduction of COBE catalyzed by these strains. Previous reports on the enzymatic reduction of COBE to (R)-CHBE with an optical purity of 92% . (Kataoka et al. 1999; Shimizu et al. 1990) recommended an organic- solvent two-phase system reaction for an enzymatic or microbial reduction, because the substrate (COBE) is unstable in an aqueous solvent and inactivates enzymes. We examined the reduction of COBE to optically pure (S)-CHBE by E. coli transformants in a water monophase system reaction and discuss the possible use of this type of reaction system in industrial applications。具有旋光性的（S）-4-氯-3-羟基丁酸乙酯在药物制剂的合成中是重要的手性化合物。其右旋体是L-卡尼汀的前体，其左旋体是羟甲基戊二酰辅酶A还原酶抑制剂的起始材料。许多研究描述了以面包酵母为基础微生物或者酶的COBE的不对称还原。我们先前已经知道利用来自念珠菌属magnoliae AKU4643 细胞催化COBE生成光学纯度96%的CHBE。这种酵母至少有三种立体选择性的还原酶，这种酵母产生的CHBE并非纯光学的，在这三种酶之中，NADPH-依赖碳酰还原酶，我们克隆并测序编码S1的基因，并在大肠杆菌中过表达。大肠杆菌转化细胞在葡萄糖，NADP+和商业化的葡萄糖脱氢酶作为辅酶因子的启动子催化COBE生成纯光学的CHBE。我们构建这三种大肠杆菌转化细胞共表达来自的S1和来自巨大芽孢杆菌的GDH，并分析COBE被这几种菌株催化还原的反应机理。先前的报道表明，利用酶催化还原COBE生成CHBE光学纯度可达92%，也提到了因为底物（COBE）在水相中不稳定，并且酶容易钝化，所以利用酶或者微生物在有机溶剂/水两相体系中催化反应。我们研究了在水单相体系中由COBE还原生成纯光学的CHBE，还讨论了这种反应体系在工业应用中可能的用途。Materials and methodsBacterial strain and plasmids The E. coli strains used in this study were JM109 and pGDA2, in which the GDH gene from B. megaterium is inserted into pKK223-3, was kindly provided by Professor I. Urabe, Osaka University (Makino et al. 1989). Plasmids pSL301 and pTrc99A were purchased from Invitrogen (USA), and Amersham Pharmacia Biotech (UK), respectively. Plasmids pUC19 and pSTV28 (Homma et al. 1995; Takahashi et al. 1995) were purchased from Takara Shuzo (Japan).材料和方法菌株和质粒本次实验中使用的大肠杆菌是JM109 and HB101。来自B. megaterium的GDH基因插入到Pkk233-3质粒中，而带有GDH基因片段的pGDA2质粒由到由大阪大学的urabe教授提供。质粒pSL301和 pTrc99A是由美国的Invitrogen公司和英国的公司分别购买的。质粒pUC19和pST28是由日本takara公司购买的。The recombinant plasmid used in this study was constructed as follows (Fig. 1): Plasmid pGDA2 was double-digested with EcoRI and PstI to isolate a DNA fragment of about kilobase pairs (kb) including the GDH gene. This fragment was inserted into the EcoRI-PstI site of plasmid pSL301 to construct plasmid pSLG. Plasmid pSLG was double-digested with EcoRI and XhoI to isolate a DNA fragment of about kb including the GDH gene.这次实验使用的重组质粒构建如下：质粒pGDA2 被EcoRI 和 PstI双酶切从而分离出一个大小约为的包含有GDH基因的DNA片段。这个片段被插入到质粒Psl301的EcoRI-PstI酶切位点从而构建出质粒pSLG。质粒pSLG被EcoRI和XhoI To construct plasmid pNTS1G, this fragment was inserted into the EcoRI-SalI site of pNTS1, which was constructed to overproduce S1 as described previously (Yasohara et al. 2000). To construct plasmid pNTGS1, plasmid pNTG was first generated. Two synthetic primers (primer 1, TAGTCCATATGTATAAAGATTTAG,and primer 2 TCTGAGAATTCTTATCCGCGTCCT) were prepared for polymerase chain reaction (PCR) using pGDA2 as the template. The PCR-generated fragment was double- digested with NdeI and EcoRI and then inserted into the NdeI EcoRI site of plasmid pUCNT, which was constructed from pUC19 and pTrc99A, as reported (Nanba et al. 1999), to obtain pNTG. To construct plasmid pNTGS1, two synthetic primers (primer 3, GCCGAATTCTAAGGAGGTTAATAATGGCTAAGAACTTCTCCAACG, and primer 4, GCGGTCGACTTAGGGAAGCGTGTAGCCACCGTC) were prepared using pUCHE, which contains the S1 gene as the template. The PCR-generated fragment was double-digested with EcoRI and SalI and then inserted into the EcoRI-SalI site of pNTG to obtain pNTGS1. Plasmid pNTS1G, pNTGS1 or pNTG was transformed into E. coli HB101.构建pNTS1是为了过表达前文所提到的S1，这个大小的片段被插入到pNTS1的EcoRI-SalI酶切位点从而构建pNTS1G。为了构建质粒pNTGS1，首先需要构建pNTG。两个合成引物（引物1，TAGTCCATATGTATAAAGATTTAG和引物2，TCTGAGAATTCTTATCCGCGTCCT）和作为模板的pGDA2是PCR反应需要的。PCR得到的片段是由NdeI 和EcoRI双酶切和并插入到质粒pUCNT的NdeI EcoRI酶切位点来得到pNTG。根据报道，pUCNT是由pUC19和 pTrc99A构建而来。为了构建质粒pNTGS1，两个合成引物(引物 3, GCCGAATTCTAAGGAGGTTAATAATGGCTAAGAACTTCTCCAACG, and 引物 4, GCGGTCGACTTAGGGAAGCGTGTAGCCACCGTC)，包括了S1基因作为模板。Pcr产物片段被EcoRI和SalI双酶切然后被插入到pntg的EcoRI-SalI酶切位点得到pntg1.质粒pNTS1G, pNTGS1或者 pNTG都是导入大肠杆菌 pGDA2 was double-digested with EcoRI and PstI to isolate a DNA fragment of about kb including the GDH gene. To construct plasmid pSTVG, this fragment was inserted into the EcoRI-PstI site of plasmid pSTV28. Plasmid pSTVG was transformed into E. coli HB101. 质粒pGDA2被EcoRI 和 PstI双酶切得到包含GDH基因的大小的DNA片段。为了构建pSTVG质粒，这个片段被插入到pSTV28质粒的EcoRI-PstI的酶切位点。pSTVG质粒被导入到E. coli HB101。Medium and cultivationThe 2×YT medium comprised Bacto-tryptone, yeastextract, and NaCl, pH . E. coli HB 101 carrying pNTS1,pNTG, pNTS1G, or pNTGS1 was inoculated into a test tube containing2 ml 2×YT medium supplemented with mg/ml ampicillin,followed by incubation at 37 °C for 15 h with reciprocal preculture ( ml) was transferred to a 500-ml shakingflask containing 100 ml 2×YT medium. The cells were cultivatedat 37 °C for 13 h with reciprocal shaking. E. coli HB101 carryingpNTS1 and pSTVG was similarly cultivated in 2×YT mediumsupplemented with mg/ml ampicillin and mg/ml chloramphenicol.培养基和培菌2*YT培养基 包含有细菌用胰蛋白胨，酵母提取物， NaCl，.携带有pNTS1,pNTG, pNTS1G, 或 pNTGS1的大肠杆菌HB101被接种到有氨苄青霉素的2ml的2*YT培养基，37°C摇床15小时。将菌液接种到100ml2*YT培养基的500ml烧瓶中。在37°C摇床培养13小时。携带有pNTS1 和 pSTVG质粒的大肠杆菌HB101在2*YT培养基中培养方法相似，只是培养基中要加入 mg/ml的氨苄青霉素和 mg/ml的氯霉素。Preparation of cell-free extracts and the enzyme assay Cells were harvested from 100 ml of culture broth by centrifugation, suspended in 50 ml of 100 mM potassium phosphate buffer (pH ), and then disrupted by ultrasonication. The cell debris was removed by centrifugation; the supernatant was recovered as the cell-free extract. Carbonyl reductase S1 activity (COBE-reducing activity) was determined spectrophotometically as follows: The assay mixture consisted of 100 mM potassium phosphate buffer (pH ), mM NADPH, and 1 mM COBE. The reactions were incubated at 30 °C and monitored for the decrease in absorbance at 340 nm. The assay mixture for GDH activity consisted of 1 M Tris-HCl buffer (pH ), 100 mM glucose, and 2 mM NADP+. The reactions were incubated at 25 °C and monitored for the increase in absorbance at 340 nm. One unit of S1 or GDH was defined as the amount catalyzing the reduction of 1 μmol NADP+ or oxidation of 1 μmol NADPH per minute, respectively. Protein concentrations were measured with a proteinassay kit containing Coomassie brilliant blue (Nacalai Tesque, Japan),using bovine serum albumin as the standard (Bradford 1976).无细胞抽提液和酶鉴定将100ml培养液离心收获菌体，用为的磷酸缓冲液悬浮，然后超声粉碎。细胞碎片通过离心可以去除，收集上层清液就是无细胞抽提物。碳酰还原酶S1的活性由分光光度计测量如下：测定的混合物包括：的磷酸二氢钾缓冲液，和1mMCOBE。反应在30°C条件下反应，并且随时监测其在340nm处的吸光值。测GDH混合物包括：1M pH 的Tris-HCl的缓冲液，100mM的葡萄糖，2mM的NADP+。反应在25°C下进行，监测其在340nm处的吸光值。一个单位S1或GDH被定义为每分钟催化还原1μmol NADP+或氧化1 μmol NADPH的量。蛋白质的测定通过含有考马斯亮蓝的蛋白质测定试剂利用牛血清白蛋白作为标准进行测定。Study of enzyme stabilityOne milliliter of 100 mM potassium phosphate buffer (pH ) containing the cell-free extracts of E. coli HB101 carrying pNTS1 (S1: 20 U/ml) was mixed with an equal volume of each test organic solvent in a closed vessel. After the mixture was shaken at 30 °C for 48 h, the remaining enzyme activities in an aqueous phase were assayed as described above. The mixture, containing 100 mM potassium phosphate buffer (pH ), S1 (20 U/ml), and various concentrations of CHBE, was incubated at 30 °C for 24 h in order to study the enzyme’s stability in the presence of remaining enzyme activities were assayed as described above.酶稳定性的研究一毫升含有含有pNTS1质粒的E. coli HB101的无细胞抽提液的100mM磷酸氢二钾缓冲液（）与等体积的有机溶剂混合。混合物在30 °C震摇48小时后，水相中残留的酶活力即是上述的酶活力。COBE reduction with E. coli cells expressing the S1 gene and E. coli cells expressing GDH genes in a two-phase system reaction The reaction mixture comprised 15 ml culture broth of E. coli HB101 carrying pNTG, 17 ml culture broth of E. coli HB101 carrying pNTS1, mg NADP+, 4 g glucose, g COBE, 25 ml n-butyl acetate, and about 25 mg Triton X-100. The pH of the reaction mixture was controlled at with 5 M sodium hydroxide. At 2 h, g COBE and g glucose were added to the reaction mixture. To compare the reaction by E. coli transformant coexpressing the GDH and S1 genes, 30 ml culture broth of E. coliHB101 carrying pNTS1G was used instead of culture broth of E. coli HB101 carrying pNTG and E. coli HB101 carrying pNTS1. Other components and the procedure were the same as described above.表达S1基因和GDH基因的大肠杆菌细胞在两相反应体系中的还原反应混合物包含有带有pNTG质粒的大肠杆菌HB101的菌液15ml，pNTS1质粒的大肠杆菌HB101的菌液17ml， mg NADP+，4 g葡萄糖，的COBE，25ml的n-butyl acetate丁酰醋酸盐和大约25mg的聚乙二醇辛基苯基醚Triton X-100。用5M的NaOH溶液将pH控制在。在反应两小时后，加入和葡萄糖到该混合物中。比较大肠杆菌转化细胞共表达GDH和S1基因，携带有pNTS1G质粒的大肠杆菌HB10130ml菌液取代了携带有pNTG和pNTS1质粒的大肠杆菌HB101菌液。其他的成分和步骤和上述的方法相似。 COBE reduction to (S)-CHBE in a two-phase system reaction The reaction mixture contained 50 ml of culture broth of an E. coli HB101 transformant, mg NADP+, 11 g glucose, 10 g COBE, 50 ml n-butyl acetate, and about 50 mg Triton X-100. The reaction mixture was stirred at 30 °C, and the pH was controlled at with 5 M sodium hydroxide. Five grams of COBE/ g glucose and 10 g COBE/11 g glucose were added to the reaction mixture at 3 h and 7 h, respectively; mg NADP+ was added at 26 在两相系统中还原生成（S）-CHBE反应混合物包含50ml E. coli HB101转化细胞的培养液，葡萄糖，10gCOBE，50ml丁酰醋酸，和大概50mg聚乙二醇辛基苯基醚Triton X-100.在30°C温度下将其混合均匀，并用5M的NaOH溶液将pH控制在。在第3小时加入5gCOBE和葡萄糖或者在第7小时加入10gCOBE和11g葡萄糖，分别在第26小时加入。 COBE reduction to (S)-CHBE in an aqueous system reaction The reaction mixture was made up of 50 ml of culture broth of an E. coli HB101 transformant, mg NADP+, 11 g glucose, and about 50 mg Triton X-100. The reaction mixture was stirred at 30 °C. Fifteen grams of COBE was fed continuously by means of a micro-feeding machine at a rate of about g/min for about 12 h. The pH of the reaction mixture was controlled at with 5 M sodium hydroxide. The reaction mixture was extracted with 100 ml ethyl acetate. The organic layer was dried over anhydrous sodium sulfate and then evaporated in vacuo. COBE在水相中还原成(S)-CHBE的反应反应的体系是由50ml大肠杆菌HB101转化细胞的菌液，，11g葡萄糖和大约50mg聚乙二醇辛基苯基醚Triton X-100。反应混合物在30°C15mg的COBE通过微量添加机器以 g/min的速率连续12小时恒定的加入到体系中。用5M的NaOH溶液将pH控制在。反应混合物用100ml乙酸乙酯萃取。有机层用无水硫酸钠吸干，并在真空中脱水。Analysis The organic layer was obtained on centrifugation of the reaction mixture and was assayed for CHBE and COBE by gas chromatography. Optical purity of CHBE was analyzed by high-performance liquid chromatography (HPLC), as described previously (Yasohara et al. 1999).Enzymes and chemicals Restriction enzymes and DNA polymerase were purchased fromTakara Shuzo (Japan). COBE (molecular weight: ) was purchasedfrom Tokyo Kasei Kogyo (Japan). Racemic CHBE (molecularweight: ) was synthesized by reduction of COBE withNaBH4. All other chemicals used were of analytical grade andcommercially available.分析离心反应混合物得到的有机层通过气相色谱法测定其CHBE和COBE。COBE的光学纯度如前所述通过高效液相色谱法进行分析。酶和化学试剂限制性内切酶和DNA聚合酶由takara公司购得，COBE（分子量：）由东京Tokyo Kasei Kogyo公司购得，消旋体CHBE（分子量）通过COBE及NaBH4合成。所有其他化学试剂都是分析等级和商业化的试剂。Construction of E. coli transformants overproducing S1 and GDHTo express the carbonyl reductase S1 and GDH genes in the same E. coli cells, four expression vectors were constructed (Fig. 1). Plasmids pNTS1G and pNTGS1 contain the S1 gene from C. magnoliae, the GDH gene from B. megaterium, the lac promoter derived from pUC19, and the terminator derived from pTrc99A. Plasmid pNTS1 contains the S1 gene, the lac promoter derived from pUC19, and the terminator derived from pTrc99A. The enzyme activities in cell-free extracts of the E. coli transformants are shown in Table 1. E. coli HB101 cells carrying the vector plasmid pUCNT had no detectable S1 or GDH activity. E. coli HB101 carrying either pNTS1G or pNTGS1 showed S1 and GDH activity without isopropyl-β-D-thiogalactopyranoside (IPTG) induction. The S1 activities of these two transformants were lower than the GDH activities. To obtain a transformant whose S1 activity was equal to or greater than the level of GDH activity, we used a lower copy vector, pSTV28 (Homma et al. 1995; Takahashi et al. 1995), to express the GDH gene. It may be possible to raise the S1 activity by lowering the GDH activity. Plasmid pSTVG contains the GDH gene, the lac promoter, the chloramphenicol resistance gene, and the replicative origin derived from pACYC184 for compatibility with the plasmid pNTS1. In E. coli HB101 carrying pNTS1 and pSTVG, the S1 activity was higher than the GDH activity, but this GDHlevel may be too low to regenerate in a COBE reduction reaction as described below.过产生S1和GDH的大肠杆菌转化细胞的构建为了在同一大肠杆菌细胞中表达碳酰还原酶S1和GDH基因，要构建四个表达型载体。质粒pNTS1G 和 pNTGS1包含有来自C. magnoliae的S1基因，来自B. megaterium的GDH基因，来自pUC19的LAC启动子，从pTrc99A的来的终止子，质粒pNTS1包含有S1基因，来自pUC19的LAC启动子，从pTrc99A的来的终止子。在大肠杆菌转化细胞的无细胞抽提物的酶活力如表一所示。携带有运输质粒pUCNT的大肠杆菌细胞无法检测到其S1和GDH活性。携带有pNTS1G 或 pNTGS1质粒在没有IPTG的诱导下有S1和GDH的活性。在这两个转化菌种中，S1的活力小于GDH的活力。为了得到S1活性等于或者大于GDH的大肠杆菌转化菌株，我们使用低拷贝的载体pSTV28，来表达GDH基因。它可能可以通过降低GDH的活性从而提高S1的活性。质粒pSTVG包含有GDH基因，lac启动子，和氯霉素抗性基因，以及与pNTS1具有相容性的从pACYC184得来的复制起始位点。在携带有pNTS1和pSTVG的大肠杆菌转化细胞中，S1的活性要高于GDH的活性，但是GDH的活性可能会太低而在COBE还原反应中不能再生。 太长了，字数有限制，所以不能发完。分数我无所谓啦，我很少登录的。这应该算是基因工程的吧，是我以前自己翻的，不是很好。如果你要的话可以联系我的邮箱。
Abstract: As the accelerating development of the molecule technology, it has been increasingly used in many grounds of human life. Along with the excited and confused feeling among people, there are definitely a large number of controversial issues on this magical technology. Especially at the Present, America attempt to prove that creating the artificial blood by genetic technology and tended to body experiment for artificial blood after 2013 has brought many arguments. This paper mainly studies the artificial blood made through the genetic technology.