5. Steerprop General_总体介绍

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欧洲药典ep8.02.6.1无菌检验sterility中英文翻译

2.6.1. STERILITY2.6.1 无菌检查法The test is applied to substances, preparations or articles which, according to the Pharmacopoeia, are required to be sterile. However,a satisfactory result only indicates that no contaminating micro-organism has been found in the sample examined in the conditions of the test.本检查方法适用于按照药典要求应当无菌的原料、制剂或其他物质。

但是，如果按照本无菌检查法的结果符合要求，仅表明在该检查条件下未发现微生物污染。

PRECAUTIONS AGAINST MICROBIAL CONTAMINATION微生物污染防范The test for sterility is carried out under aseptic conditions.In order to achieve such conditions, the test environment has to be adapted to the way in which the sterility test is performed. The precautions taken to avoid contamination are such that they do not affect any micro-organisms which are to be revealed in the test. The working conditions in which the tests are performed are monitored regularly by appropriate sampling of the working area and by carrying out appropriate controls.无菌检测试验应在无菌的条件下进行。

Plasmid Premier中文使用说明书

Plasmid Premier 2.02Plasmid Premier是由加拿大的Premier Biosoft公司推出的用于质粒作图的专业软件，主要用于进行质粒作图，质粒特征分析和质粒设计。

其主要界面分为序列编辑窗口（Genetank），质粒作图窗口（Plasmid Design），酶切分析窗口（Restriction Sites）和纹基分析窗口（Motif）。

打开程序就进入以下的序列编辑窗口，可以直接打开Genbank或Vector数据库中已知质粒的序列文件，将序列读入，并将有关于质粒的各种特征，包括编码区，启动子，多克隆位点以及参考文献等信息保存在Header中；也可以直接输入序列进行未知质粒的设计。

在该窗口中显示质粒序列的方式可以有按照正向链，反向链和双链格式显示，并在显示碱基分组上选择3个/组或10个/组，以及用5’ Seq No命令选择在环形质粒中开始编辑的序列5’端位置。

该程序提供的翻译功能是该程序独到的地方，可以实现DNA，RNA和蛋白序列之间的互相翻译，并且提供目前已知的所有密码表来进行翻译，还能够是用户可以对密码表进行编辑，有很高的自由度。

最为重要的是该程序提供了由蛋白向DNA按照IUPAC密码表进行逆向翻译的功能，得到了Translated DNA序列，通过在该序列中寻找序列中简并性低的区段由于进行猜测体探针的设计，这是该程序提供的在质粒作图之外的一个比较有特点的用途。

在序列编辑窗口中可以通过点击按钮进入以下的质粒作图界面在质粒图谱中主要包括质粒的四种特征元素，包括限制性酶切位点，特征序列（纹基，motif），开读框（ORF）和其他质粒特征（feature如编码区）。

其中前三种元素都是直接由程序对序列直接分析获得，也可以由用户再通过analyze 菜单再作进一步分析，而最后一中元素可以由header头信息定义，也可以由用户自由添加。

并切均可获得四种元素的列表。

对于图谱中各种元素，Plasmid Premier提供良好的图象处理支持，可以自由拖拽图中显示的各个元素，改变各个元素的显示，包括各个元素的字体，名称，位置和色彩等，并且通过点击途中的各个元素来获得其在序列上的相应位置，并且可以通过选择按钮选择只显示其中的一种元素，使界面简洁，易于进行编辑处理。

minitab 在质量管理中的英语-培训笔记

1.Pareto图得出关键少数（找出众多问题中的主要问题，优先排钱）2.用因果（鱼骨）图分析主要问题的原因（观点），末端原因应能一眼看出对策（员工未按标准作业执行，不能成为末端原因）3.用散点图（采集数据大于30个）验证因果是否具有相关性4.直方图（采集数据大于30个）显示数据频度分布，拟合的曲线为正态分布曲线分布的均值：衡量数据的准确性标准差：数据的精确性（标准差越小，精确度越好）6倍标准差（包含99.73%的数据）=过程能力（身高差超过6*15.47的可能性很小）5. 单值图，箱线图，时间序列图6.测量系统分析（需增加2张手机拍的图）A．交叉（不破坏）此图只具有参考意义测量值*测量员体现重复性来源标准差(SD) (6 * SD) 异 (%SV) (SV/Toler)合计量具 R&R 0.067596 0.40558 33.56 40.56（两者都小于10%，合格；10%~30%测量关键特性，任意一个大于30%，不合格）重复性 0.032592 0.19555 16.18 19.56再现性 0.059220 0.35532 29.40 35.53（再现性影响更大）测量员 0.028470 0.17082 14.13 17.08测量员*洗衣粉袋 0.051928 0.31157 25.78 31.16部件间 0.189745 1.13847 94.20 113.85合计变异 0.201426 1.20856 100.00 120.86产品过程可区分的类别数 = 3（可区分类别数>10,优秀；大于等于5，不合格；小于5，不及格）（衡量分辨力）综上红色字体，以上测量系统不合格B.嵌套（破坏性）嵌套式与交叉式相比，缺少再现性图过程公差 = 16研究变异 %研究变 %公差来源标准差(SD) (6 * SD) 异 (%SV) (SV/Toler)合计量具 R&R 1.59164 9.5499 42.86 59.69重复性 1.59164 9.5499 42.86 59.69 再现性 0.00000 0.0000 0.00 0.00 部件间 3.35534 20.1321 90.35 125.83 合计变异 3.71371 22.2823 100.00 139.26可区分的类别数 = 2分析同上，测量系统不合格7.测量线性研究偏倚——点线性——计数型测量系统分析评定值的属性一致性分析检验员自身（重复性）评估一致性# 检 # 相检验员验数符数百分比 95 % 置信区间钱 6 3 50.00 (11.81, 88.19)孙 6 4 66.67 (22.28, 95.67)赵 6 6 100.00 (60.70, 100.00)# 相符数: 检验员在多个试验之间，他/她自身标准一致。

5. Steerprop General_总体介绍

Steerprop LTD is the center of azimuth propulsion technology. Steerprop Ltd. produces azimuth propulsors for maritime and offshore industries. Steerprop公司，是以全回转推进器技术为中心，为船舶和海洋工程提供产品。

We provide the customers with outstanding quality and excellent lifetime economy. Steerprop operates on a global basis by providing azimuth main propulsion solutions for the high seas, as well as coastal and inland waters everywhere in the world.Steerprop向顾客提供高品质、寿命长的产品。

以国际化的标准，致力于提供有关远洋全回转推进器相关问题的解决方案。

The mission of Steerprop Ltd. is to equip all eligible vessels with azimuth propulsors and maintain them together with the co‐operation partner networks. According to the company’s vision, Steerprop Ltd. is to become the leading technological expert within the azimuth propulsion industry. Steerprop公司的宗旨，是使所有合适的船只全部装配上全回转推进器并且与他们形成合伙供应商网络。

表型组精密测量术语

表型组精密测量术语表型组精密测量术语是指在研究生物个体表型（即外部可观察的性状或表现）时，使用的精密测量技术和相关术语。

以下是一些常见的表型组精密测量术语：1. 全基因组测序（Whole genome sequencing）：对个体的完整基因组进行测序，包括所有基因和非编码DNA序列的测量。

2. 转录组测序（RNA-Seq）：对个体的基因组中转录的RNA分子进行测序，以了解基因表达的水平和差异。

3. 蛋白质组测序（Proteomics）：对个体中蛋白质的组成、结构和功能进行全面的测量。

4. 代谢组测序（Metabolomics）：对个体中代谢产物（代谢物）的组成和变化进行精密测量，以了解代谢途径和生物化学反应。

5. 表型测量（Phenotyping）：对个体的特定性状进行定量和定性测量，包括外部形态、生理特征、行为特征等。

6. 高通量测量（High-throughput measurement）：使用自动化和高通量技术对大量样本进行快速测量的方法，例如基因芯片、流式细胞术等。

7. 生物图像学（Biological imaging）：使用显微镜、成像仪等设备对个体的形态、结构和功能进行图像化测量。

8. 多样本分析（Multi-sample analysis）：对多个样本进行同时分析和比较，以揭示群体间的差异和关联。

9. 数据挖掘和统计分析（Data mining and statistical analysis）：对测量数据进行处理、分析和解释，寻找关键特征、趋势和关联。

这些术语涵盖了一系列不同的测量技术和分析方法，可以帮助科学家更深入地研究个体表型的多个方面，从而揭示基因型与表型之间的关系和相互作用。

新波特公司 TSP 阶段系列精确线性阶梯说明说明书

WarrantyNewport Corporation warrants its product to be free of defects in material and workman-ship for a period of twelve (12) months from the date of shipment. During the warranty period, Newport will repair or replace (at Newport’s discretion) any component within its product that fails to adhere to published specifications (for standard product) or specifi-cations in the quotation (for custom/special product).For warranty service requiring return of a product to Newport, the item(s) must be re-turned to our Service Center, with all shipping, taxes, or duty charges prepaid unless special arrangements have been approved beforehand by Newport. The location of services performed under warranty will be determined by Newport.If using controller electronics other than Newport’s, the liability of the product rests with the customer if upon performance evaluation of the stage there are no anomalies exhib-ited with Newport’s electronics.Warranty LimitationsThe foregoing warranty shall not apply to defects resulting from:ponents and accessories manufactured by companies other than Newport,which causes damage to Newport’s products, nor does Newport’s warranty coverthe cost of the customer’s time and expenses incurred in diagnosing, repairingand handling Newport’s warranty related issues.b.Improper or inadequate maintenance by the buyer not in adherence withNewport’s guidelines.c.Customer-supplied interfacing.d.Operation outside the environmental specifications of the product.e.System malfunctions that are related to or caused by software misuse.f.Improper site preparation and mainenance of unauthorized product modificationor misuse.g.Warranty void if not delivered by an authorized Newport Corp distributor.Newport assumes no liability for customer-supplied material. The obligations ofNewport are limited to repairing or replacing, without charge, equipment whichproves to be defective during the warranty period only. The warranty on partspurchased after the expiration of the original warranty is ninety (90) days. Ourwarranty does not cover damages due to misuse, negligence or accidents, ordamages due to installation, repairs or adjustments not specifically authorizedby Newport Corp.STATEMENT OF CALIBRATIONThis instrument has been inspected and tested in accordance with specifications published by Newport Corp.The accuracy and calibration of this instrument (where applicable) is traceable to the National Institute for Standards and Technology through equipment which is calibrated at planned inter-vals by comparison to the certified standards maintained at Newport Corp.ACCURACY AND REVISIONSThe information in this document has been checked and is believed to be entirely reliable.However, no responsibility is assumed for inaccuracies or inadvertent omissions. Furthermore, Newport Corp. reserves the right to make changes to any product herin to improve reliability, function or design.USE OF THIS MANUALPlease read through this manual carefully before using the equipment provided. It should benoted that if the equipment is used in a manner other than that specified by this manual, per-sonal injury or damage to the equipment may be incurred.SAFETY CONSIDERATIONSHazard Warning LabelsThe hazard warning labels pictured below are affixed to some Newport equipment. In Figure 1 below, is a Pinch Point warning label. This indicates that injury could occur if extremities areplaced between moving components.Figure 1Figure 2Abbildung 2. ElektroshockgefahrFigura 2. Riesgo EléctricoFigue 2. Risque électriqueDisconnect power to motorized equipment under the following circumstances:AchtungDie Stromversorgung motorbetriebener Geräte is in folgenden Situationen zu unterbrechen:· Wenn das Gerät Regen oder übermäßigerFeuchtigkeit ausgesetzt ist,oder wenn Flüssigkeiten in das Gerät geraten sind.· Wenn das Gerät fallengelassen wurde oder das Gehäuse beschädigt ist.· Wenn dasangenommen wird, daß Service - oder Reparaturarbeiten erforderlich sind· Wenn das Gerät gereinigt wird.AdvertenciaDesconecte el equipo motorizado de la alimentación CC bajo las siguientes circunstancias:·Si se expone la unidad alluvia, humedad excesiva o si le cae encim algún líquido.· Si la unidad se ha caído o se ha dañado su alojamiento.· Si se sospecha que requiere mantenimiento o reparción.· Cada vez que se limpie el alojamiento.AttentionDébranchez l'alimentation secteur de l'appareil de pilotage dans les cas suivants:· Si l'équipement a été exposé à la pluie, à une humidité excessive ou à des projections de liquide.· Si l'équipement a subit un dommage durant le transport(chute ou emballage abîmé).· Si vous estimez que l'équipement a besoin d'une réparation.· Chaque fois que vous nettoyez l’équipement.To avoid hazardous situations and to protect the equipment from danger, observe the following recommendations:Do not make any modifications or parts substitutions to the units.Do not touch, directly or with other objects, live circuits inside motorized units.Do not operate the units in an explosive atmosphere.Do not expose the units to excessive moisture (>75% relative humidity).UNPACKING AND INSPECTIONAll Newport products are carefully assembled, tested and inspected before shipment.Upon receiving this product, please check for any obvious signs of physical damagethat might have occurred during shipment. Report any such damage to the shippingagent immediately. Retain the original packaging materials in case reshipmentbecomes necessary.If a Newport product must be returned, the following information is needed for proper shipment back to Newport:a. Unit Model Numberb. Unit Serial Numberc. Reason for return.A Return Material Authorization number (RMA) will be issued, which should be referenced onyour shipping documents. Please fill out the included service form and return it with the unit.Use the proper precautions when shipping the unit. Damage incurred during shipping mayinvalidate your warranty. Please contact our technical support or returns group at 800-222-6440 for more information.SET UPRemoving Shipping Tabs (Figure 3)To prevent damage during shipping, the top plate of the stage has been locked with red anod-ized tabs. Orange warning tags indicate the position of each tab. To remove the tabs, undertake the following steps:e 5mm hex wrench to remove socket head cap screws (M6X1) from each lock tab formetric stages, or 3/16 hex wrench to remove socket head cap screws (1/4-20) fromeach lock tab for english stages.2.Remove the lock tab and warning tag and retain for future relocation.Figure 3Environmental ConsiderationsTemperatureFor optimum performance, the stage should be maintained at a temperature of20o C (68o F +/- 2o F).VibrationThe stage should not be set up within the vicinity of vibration or shock. Outside forces, such as forklifts, compressors and other machinery, can induce unwanted motion in the stages via the floor on which the whole system is resting. Resulting vibrations can lead to measurement errors.Surface Plate FlatnessTo ensure accuracy, the surface plate on which the stage is mounted should be flat to within2.5um (0.0001 inch) over all. Granite is the preferred mounting surface.Fastener KitA fastener kit containing screws, and flat washers, are provided for bolting the stage to a sur-face plate. The kit also contains anti-pinch caps for plugging unused slots.Table 1ScrewM6-1 x 35mm 1/4-20 x 1 3/8inch M6inside diameterWasher MetricEnglishStage InstallationAllow clearance for the 25 pin connector.Bolting the Stage to the Surface Plate1.Clean the surface plate prior to mounting the stage.2.The stage should be attached at all four corners to the surface plate. To allow access tothe counter bores, move the stage alternately to one end-of-travel limit and then to theother. Use fastener kit (A00000xxx) provided. The screws should be torqued to between13-14.7 N-m (115-130 in-lbs).3.After torquing the fasteners, place the supplied anti-pinch caps into the mounting holes.Ensure that the caps do not protrude above the surface of the bottom stage plate. Use of these caps is mandatory to ensure CE compliance.Figure 44.Connect the cable from the stage to the motion contoller using the 25 pin interconnectcable supplied.Initial OperationThe TSP Stage has been adjusted and calibrated at the factory, prior to shipment. Under normal circumstances, no calibrations or adjustments are required after the equipment has been set up.However, it is highly recommended that initial verifications of proper limit switch operation and of proper configuration of the controller are performed.Note: If using a controller other than a Newport controller, the stage may not work properly.Contact Newport technical assistance for modification instructions.Verification of Proper Limit Switch OperationThe cable needs to be connected and the controller under power to perform this test. When the controller is referenced below, refer to the manufacturer’s manual for the controller (and control-ler software, if in use) to find the relevant procedures.1.Disconnect power to the motor via the controller.ing the manual knob, move the stage to the counterclockwise end-of-travel limit.3.Check the status of the limit via the controller.4.If the controller does not indicate arrival at the limit, refer to the Home and LimitSwitches Troubleshooting guide.5.Repeat steps 3 to 5 for the clockwise end-of-travel limit.Verification of Proper Controller ConfigurationBefore using the stage for any application, the controller should be specifically configured and programmed for operation with the TSP stage. Refer to the manufacturer’s manual for the controller to find the relevant procedures. Fine tuning of the controller may also be required. 1.Program the controller so that the first move of the stage is a short distance, low velocitymove. The move should be a distance of less than a quarter of full travel at a velocity of less than 2 mm/sec (.01 in/sec).2.Perform the first move. Use a suitable indicator to check that the stage has travelled therequired distance.3.If the stage has not travelled the required distance, refer to the Stage and ControllerTroubleshooting Guide.Attaching to the StageComponents’ Surface FlatnessThe mating surfaces of components being attached to the stage should be flat to within 2.5um (0.0001 inch) overall.Screw SpecificationsUse an M6X1 or 1/4-20 screw, depending on whether the mounting holes are metric or english to attach other components. The length of the screw protruding into the stage should be no more than 12mm (1/2 inch).TorqueComponents being attached to the stage should be torqued in an alternating pattern. The maximum torque should not exceed 11.3 N-m (100 in-lbs).MomentComponents being attached to the stage should not induce a moment of greater than 22.6 N-m (200 in-lbs).Locations of the connector, end-of-travel (CCW) and (CW), circuit board, home sensor, and ball screw.Connector Circuit Board End Limt(CW)BallscrewHome SensorEnd Limit (CCW)TSP 300 ShownMAINTENANCEThe frequency of required cleaning, lubrication and testing is highly user dependent. Refer to Table 2 below to establish a maintenance schedule for your unit.Table 2LubricationWhen applying lubricant, do not use either a harsh applicator which may scratch or damage the stage surfaces, or a fragile applicator, which may generate particles to contaminate the compo-nent. The recommended lubricant is Lamora® D68 Oil.Figure 91.Inspect the bearing rails for signs of damage or corrosion. Consult Newport’s technicalassistance group if damage or corrosion has occured.e the manual knob to move the stage to one end-of-travel limit.3.Clean the exposed bearing rail v-grooves on both upper and lower rails with Isopropylalcohol and a lint free cloth.4.Apply a light film of Lamora® D68 oil to the exposed v-grooves and to the top of the rail(Figure 9).5.Move the stage to the other end-of-travel limit and repeat steps 2 and 3.6.Run the stage from one end-of-travel limit to the other to distribute the lubricant evenlyalong the rails.Ballscrew LubricationThe recommended lubricant for the ball screw is NBU8 EP grease.Figure 10e a 2 mm hex wrench to remove the flat head cap screws from the drivecover. Remove the drive cover.2.Inspect the ballscrew for signs of damage or wear.3.Clean the ballscrew with a lint free cloth, and isopropyl alcohol. Manually move the stageas required to gain access to all of the ballscrew.4.Apply light file of the lubricant to the exposed part of the ballscrew (Figure 10).5.Manually run the stage from one end-of-travel limit to the other to distribute the lubricantevenly along the ballscrew.6.Wipe off any access lubricant with a lent free cloth.7.Replace the drive cover.DIMENSIONS(SEE CHARTS ON NEXT PAGE)Linear EncodersNon-Contact Encoder.1 um ResolutionTTL Quad AB Output (Differential)4 um Signal PeriodHome (Origin) at negative end of travelSTAGE AND CONTROLLER TROUBLESHOOTING GUIDEprovided. The various problems along with the possible causes and corrective actions required are outlined below.This section may help you to troubleshoot common problems with operating the equipment ProblemStage does not functionStage moves a short distance and stops. Controller reports “Following Error” or other motion error. Stage motion un-steady or jerkyStage travels less than specifications Stage does not meet specificationsTighten connector.Check that cables are correctly connectedand wired.Refer to the manufacturer’s manual for thecontrollerRefer to the manufacturer’s manual for thecontrollerCheck for and remove all obstructionsTighten couplingTighten connectorMotor plus/minus leads reversed and/ortachometer plus/minus leads reversedContact Newport for assistanceLower velocity and/or accelerationCheck for and remove all obstructions. Cleanrails according to the procedures outlined inthis manualTighten connectors. Check cabling andreplace or repair as necessaryContact Newport for assistanceRefer to the manufacturer’s manual for thecontrollerCheck for and remove all obstructionsEnsure that all cables are clear of the movingequipmentIf roller creep is suspected, contact Newportfor assistanceRefer to Setup, Environmental consider-ations in this manualRefer to Setup, Environmental consider-ations in this manualRefer to the specifications in this manual andconsult Newport for assistanceContact Newport for assistanceConsult Newport for recalibration or certifica-tionLoose connector at stageStage incorrectly connected to control-lerController incorrectly programmed.Limits not enabled via softwareController not connected to powersourceObstruction to movementLoose motor couplingLoose connectors at stage or controllerIncorrectly wired cablingStage incorrectly tunedLoad too large for motion profile usedObstrucion or foreign object on bearingrailsLoose connections or faulty cablingStage incorrectly tunedController incorrectly programmedObstruction to motionEntangled cablesRoller creepImproper mountingImproper attachmentStage outside of environmental specifi-cationsStage incorrectly tunedImproper handling and/or transportCause Corrective ActionStage makes un-usual noise Controller reports that both end limits are active Object contacting the manual operationknobLoose coversEntangled cablesStage incorrectly tunedLoose connectorFaulty or miswired cablesController incorrectly programmedRemove objectTighten screwsEnsure that all cables are clear of the mov-ing equipmentContact Newport for assistanceTighten all connectorsCheck cabling and rewire or repair as neces-saryRefer to the manufacturer’s manual for thecontrollerProblem Cause Corrective ActionHOME & LIMIT SWITCHES TROUBLESHOOTING GUIDETransmissive Sensor TestingPerform this procedure for the following sensors:•Clockwise•Counterclockwise•Home1.Find a thin strip of flexible material, such as heavy paper or thin cardboard, to use as aflag.2.Disconnect power to the stage.3.Remove the four flathead cap screws from the drive cover. Remove the drive cover.4.Manually position the stage to allow access to the sensor under test.mand the controller to disable motor power. Refer to the manufacturer’s manual forthe controller to find the relevant procedure.6.Reconnect power to the stage.7.Insert the flag between the two upright posts of the sensor. The sensor shouldswitch off when the strip is inserted and illuminate again when the strip is removed.SensorClockwise limitCounterclockwise limitHomePINOUT CONNECTIONSThe circuit board connector interfaces to the motion controller. Refer to the below pinout chart for pinout numbers.Pin No.12345678910111213141516171819202122232425DC Servo Motor + Tachometer Generator+ Tachometer Generator- Tachometer Generator- Tachometer Generator+ Motor Phase+ Motor Phase- Motor Phase- Motor PhaseNo ConnectionNo ConnectionNo ConnectionNo ConnectionHome Switch Signal Shield GroundEncoder Index Pulse I Limit Ground+ Travel Limit- Travel LimitEncoder Channel A Encoder Channel B Encoder Power +5V Encoder GroundEncoder Channel /A Encoder Channel /B Encoder Index Pulse /I。

PCR Primer Design

Molecular Biology Today 2001. 2(2): 27-32.P C R P r i m e r D e s i g nVinay K. Singh and Anil KumarBioinformatics Sub-centre, School of Biotechnology, Devi Ahilya University, Khandwa Road, Indore 452 017 MP, IndiaAbstractTo make PCR a specific, efficient and cost effective tool for researchers and clinicians the most important aspect is oligonucleotide primer design. This review discusses various aspects of primer design. Advice is provided for optimal design and the role of bioinformatic tools is highlighted. The authors discuss theoretical considerations and compare computational and experimental studies.IntroductionBioinformatics is a newly-emerged inter-disciplinary research area spanning a range of specialties that include molecular biology, biophysics, computer science, mathematics and statistics. It makes use of scientific and technological advances in the areas of computer science, informationtechnology and communication technology to solve complex problems in life sciences, particularly problems in biotechnology. Bioinformatics comprises of the development and application of algorithms for the analysis and interpretation of data, for the design and construction of vital databases, and for the design of experiments.Bioinformatics is used interchangeably with the terms biocomputing and computational biology. However, biocomputing is more correctly defined as the systematic development and application of computing systems and computational solution techniques to model biological phenomena. Polymerase chain reaction (PCR) is one such phenomenon. PCR is used for the in vitro生物秀-专心做生物w w w .b b i o o .c o mamplification of DNA at the logarithmic scale. Various components of the PCR reaction such as Taq DNA polymerase, assay buffer, deoxynucleoside triphosphates, stabilizing agents, and primers make it possible for the DNA template to be amplified sufficiently in vitro to attain detectable quantities. PCR can be used for various purposes such as the amplification of human specific DNA sequences, differentiation of species, sub-species and strains, DNA sequencing, detection of mutations, monitoring cancer therapy, detection of bacterial and viral infections, pre-determination of sex, linkage analysis using single sperm cells, ascertaining recombinant clones and studying molecular evolution. PCR is a sensitive technique and therefore highly susceptible to contamination which may result in false positivity. To make PCR a specific, efficient and cost effective tool for researchers and clinicians the most important component of the PCR is the oligonucleotide primers. Literature searches indicate that insufficient experimental work has been done in the field of bioinformatics especially in the field of nucleic acid sequence analyses. Inadequate experimental data is available (at least in the public domain) for the establishment of primer design strategies. In this review the authors aim to establish various aspects and types of PCR and primer design theory, supported by computational and experimental data.PCR Primer DesignSelective amplification of nucleic acid molecules, that are initially present in minute quantities, provides a powerful tool for analyzing nucleic acids (Saiki et al., 1985; Mullis et al ., 1987). The polymerase chain reaction is an enzymatic reaction, which follows relatively simple, predictable and well understood mathematical principles. However the scientist often relies on intuition to optimise the reaction. To make PCR an efficient and cost effective tool, some components of PCR such as Taq DNA polymerase, assay buffer, deoxynucleoside triphosphates (dNTPs), stabilizing agents (Sarkar et al ., 1990), DNA Template and oligonucleotide primers must be considered ingreater detail (Linz et al., 1990). Efficacy and sensitivity of PCR largely depend on the efficiency of primers (He et al., 1994). The ability for an oligonucleotide to serve as a primer for PCR is dependent on several factors including: a) the kinetics of association and dissociation ofprimer-template duplexes at the annealing and extension temperatures; b) duplex stability of mismatched nucleotides and their location; and c) the efficiency with which the polymerase can recognize and extend a mismatched duplex. The primers which are unique for the target sequence to be amplified should fulfill certain criteria such as primer length, GC%, annealing and melting temperature, 5' end stability, 3' end specificity etc (Dieffenbach et al., 1993).DNA template quality or purity is not particularly significant for amplification. Provided DNA does not contain any inhibitor of Taq DNA polymerase, it can be isolated by almost any method (Murray and Thompson, 1980; Sambrook et al ., 1989; Kaneko et al., 1989; Mercier et al., 1990; Kawasaki 1990a; Green et al., 1991; Keller et al., 1993; Klebe et al., 1996; Singh and Naik, 2000). Taq DNA polymerase also plays an important role (Drummond and Gelfand, 1989). Taq DNA polymerase from different suppliers may behave differently because of the different formulations, 生物秀-专心做生物w w w .b b i o o .c o massay conditions and/or unit definitions. Recommended concentration ranges between 1-2.5 Units/50-100 ml reaction (Lawyer et al.,1989) when other parameters are optimal.Most of the reviews on PCR optimization (Erlich et al., 1991; Dieffenbach 1993; Roux 1995) consider different parameters of PCR but generally do not discuss basic concepts of PCR primer design. Because of the requirements for different strategies of PCR, more effective PCR studies would be attainable by considering the basic concepts of PCR primer design.Primer Length: a Hard Core FactorLength of a primer is a critical parameter (Wu et al., 1991). The rule-of-thumb is to use a primer with a minimal length that ensures a denaturation temperature of 55-56°C. This greatly enhances specificity and efficiency. For general studies, primers of typically 17-34 nucleotides in length are the best. Primer >16 nucleotides in length are not generally annealed specifically to non-target DNA sequence (e.g. human DNA in an assay for bacterial infection). For example, a short primer sequence, such as a 12 bp oligonucleotide, binds to 200 specific annealing sites in the human genome. (The genome consists of 3x109 nucleotides: 3 x 109/412=200). In contrast, a 20 mersequence is expected to randomly exist only once every 420 nucleotides and as such, has only a 1 in 400 probability of existing by chance in the human genome. Primers, 18-24 mer are accepted as best in being sequence specific if the annealing temperature of the PCR reactions is set within 5°C of the primer T d (dissociation temperature of the primer/template duplex) (Dieffenbach, 1993).Primers work exceptionally well for the sequence with least intra-strand secondary structure. This is because secondary structure impedes primer annealing and extension. Longer primers (28-35 mer) are required only to discriminate homologous genes within different species or when a perfect complementary sequence to all the template is not expected. They could also be used when extra sequence information e.g. a motif binding site, restriction endonuclease site or GC clamp is attached to 5' end. Such extensions do not generally alter annealing to the sequence specific portion of the primer (Sheffield et al., 1989).Although the following formula is generally used for determining melting temperature (Tm): Tm = 4 (G+C) + 2(A+T)Frier et al. (1986) showed that the nearest-neighbor calculation is better for calculating the melting temperature of longer primers because this also takes account of thermodynamic parameters. Using improved nearest-neighbor thermodynamic values given by John SantaLucia (1995), a good estimate of melting temperature can be obtained for oligonucleotide analysis.生物秀-专心做生物w w w .b b i o o .c o mTerminal Nucleotides Make a DifferenceBoth the terminals of the primer are of vital importance for a successful amplification. The 3'-end position in the primer affects mispriming. However, for certain reactions, such as amplification refractory mutation system (ARMS), this mispriming is required (Newton et al., 1989; Old, 1991; Tan et al., 1994). Runs (3 or more) of C's or G's at the 3' end of the primer should be avoided as G +C rich sequence leads to mispriming. Complementarity at the 3' end of the primer elevatesmispriming as this promotes the formation of a primer dimer artifact and reduces the yield of the desired product (Huang et al., 1992). The stability of the primer is determined by its false priming efficiency; ideally it should have a stable 5' end and an unstable 3' end. If the primer has a stable 3' end, it will bind to a site which is complementary to the sequence rather than the target site and may lead to secondary bands. It is adequate to have G or C in last 3 bases at 5' termini for the efficient binding of the primer to the target site. This GC clamp reduces spurious secondary bands (Sheffield et al., 1989).GC Content, Tm and Ta are InterrelatedGC content, melting temperature and annealing temperature are strictly dependent on one another (Rychlik et al., 1990). GC% is an important characteristic of DNA and provides information about the strength of annealing. A GC of 50-60% is recommended. The value recommended by Dieffenbach (1993) is 45-55%.Secondary StructureAn important factor to consider when designing a primer is the presence of secondary structures. This greatly reduces the number of primer molecules available for bonding in the reaction. The presence of hairpin loops reduces the efficiency by limiting the ability to bind to the target site (Singh et al., 2000). It is well established that under a given set of conditions, the relative stability of a DNA duplex structure depends on its nucleotide sequence (Cantor and Schimmel, 1980). More specifically, the stability of a DNA duplex appears to depend primarily on the identity of the nearest-neighbor nucleotides. The overall stability and the melting behavior of any DNA duplex structure can be predicted from its primary sequence if the relative stability (DG 0) and thetemperature dependent behavior (DH 0, DCp 0) of each DNA's nearest-neighbor interaction is known (Marky and Breslauer, 1982). Tinoco et al., (1971, 1973) and Uhlenbeck et al., (1973) have predicted stability and melting behavior of RNA molecules for which they and others have 生物秀-专心做生物w w w .b b i o o .c o mdetermined the appropriate thermodynamic data. But, to the best of our knowledge, no experimental data is available to support the prediction of the thermodynamic properties of hairpin structures, an important factor to consider when designing a primer. Single stranded nucleic acid sequences may have secondary structures due to the presence of complementary sequences within the primer length e.g. hairpin loops and primer-dimer structures. We have recently shown experimentally that hairpin loops, if present, can greatly reduce the efficiency of the reaction by limiting primer availability and the ability to bind to the target site (Singh et al., 2000). The effect of primer-template mismatches on the PCR has been studied earlier in a Human Immunodeficiency Virus (HIV) model (Kwok et al., 1990). Studies have also been performed for the characterization of hairpins (Marky et al., 1983, 1985), cruciforms (Marky et al., 1985), bulge and interior loops (Patel et al., 1982 , 1983).Dimers and False Priming Cause Misleading ResultsAnnealing between the 3' end of one primer molecule and the 5' end of another primer molecule and subsequent extension results in a sharp background product known as primer dimer. Its subsequent amplification product can compete with the amplification of the larger target. If the primer binds anywhere else than the target site, the amplification specificity is reduced significantly (Breslauer et al., 1986). This leads to a weak output or a smear. This occurs again when some bases at 3' end of the primer bind to target sequence and achieve favorable chances of extension (Chou et al., 1992). To minimize the possibility of dimers and false priming, PCR is generally performed at hightemperature (>50°C), but primers may be extended non-specifically prior to thermal cycling if the sample is completely mixed at room temperature (RT) (Hung et al., 1990). To prevent this occurring the Hot Start ® protocol is recommended (Erlich et al., 1991). All reagents except one (usually the Taq DNA Polymerase) are mixed at RT. The sample is denatured completely for 3 to 7 min, kept on ice for 2 min and then Taq DNA polymerase is added to start the reaction.Know Your Product Before AmplificationPCR product length is directly proportional to inefficiency of amplification (Wu et al., 1991). Primers should be designed so that only small regions of DNA (150-1000 bp) can be amplified from fixed tissue samples or purified plasmid or genomic DNA. The product is ideal for probe hybridization studies (Schowalter and Sommer, 1989). For reverse transcriptase polymerase chain reaction (RT-PCR) as described by Kawasaki (1990b), primers should only be designed in exons taking care that both primers should be on different exons of mRNA to avoid spurious product amplified from contaminating DNA in the mRNA preparation, if any. If the desired restriciton 生物秀-专心做生物w w w .b b i o o .c o menzyme site is not available within the amplified product, it may be incorporated within the primer (Ponce and Michal, 1989; Jung et al., 1990).Mismatch to Improve Sensitivity and SpecificityThere is a good and a bad aspect to mismatches in primers. Single mismatches at or near theterminal 3' nucleotide of a primer are known to affect both oligonucleotide stability and efficiency of polymerase reaction; mismatches in the primer at or near the 3' terminal end affect PCR more dramatically than mismatches at other positions (Petruska et al., 1988). Generally, mismatches at the 3' end terminal nucleotide reduce or inhibit efficiency of amplification (Kwok et al., 1990; Liu et al., 1994) but studies have shown that a mismatch 3-4 bases upstream of the 3' end of a primer used for the ARMS study actually increases specificity. A mismatch may therefore be deliberately created while designing a primer for ARMS PCR (Old, 1991).Nested PCRNested PCR is often successful in reducing unwanted products while dramatically increasingsensitivity (Albert and Fenyo, 1990). It is used when the actual quantity of target DNA is very low or when the target DNA is impure. Nested PCR reduces background amplification thereby enhancing target detection. The technique is especially helpful for amplification of low copy number targets (<100 molecules) and while doing quantitative PCR (Haff, 1994). The process involves one PCR reaction followed by the next PCR extension which amplifies the first PCR product. Two sets of primers are designed. PCR is first carried out using outer primers andsubsequently with inner primers positioned within the product obtained in the first extension. It is also possible to perform a nested PCR reaction in a single sample without dilution between the two PCR reactions (Erlich et al., 1991). When designing primers for nested PCR, care must be taken to eliminate potential primer dimers and cross dimers within and between inner and outer primer sets.Multiplex PCRThis technique involves co-amplification of two or more target sequences within a single sample (Chamberlain et al., 1991; Edwards and Gibbs, 1994). A unique pair of primers for each target is preferred but primers can be designed so that a single primer can amplify different regions with two or more counterparts (Varawalla et al., 1991a; 1991b). When designing primers for multiplex PCR 生物秀-专心做生物w w w .b b i o o .c o msystems, the basic rule is to have similar annealing temperatures and similar GC% of the primers (Nicodeme and Steyaret, 1997). Product length should also be taken into consideration when designing primers so that they can be effectively separated and studied by electrophoresis.Multiplex PCR may be used for detection of genetic disorders (Old et al., 1990; Shuber et al., 1991). Zhu and Clark (1996) demonstrated that addition of competitive primers may dramatically increase PCR amplification efficiency.Universal PrimersMolecular biologists are well aware of the exponential increase in the DNA sequence databanks with several thousands bases added every day. Many genes of varied importance have beensequenced in several species. However, the scientific community may require information on such genes in other species, which are used as experimental models. Researchers are often forced to re-sequence genes for new species in order to conduct expression level or other PCR related studies of the gene (Kain et al., 1991) Bulat et al. (1992) demonstrated the application of universal primers. Universal primers facilitated the rapid study of novel genes in new models. Rose et al., (1998) demonstrated a new primer design strategy for PCR amplification of unknown targets that are related to multiple-aligned protein sequences. Universal primers are designed in the conservedregion of the sequences (Singh et al., 2000). Universal primers should be designed from amino-acid sequences in the regions of lowest degeneracy using a multiple sequence alignment (Nomenclature Committee of the International Union of Biochemistry, 1985). Universal primers were used for differential display of eukaryotic mRNAs by PCR (Liang and Pardee, 1992). A universal primer set for detection of parasitic genomes was also designed using Dirofilaria immitis as a test sample (Nagano et al., 1996), whereas Venta et al. (1996) designed gene-specific universal primers for the canine genome. These were used for developing a genetic map of dog-based markers. Universal primers may be used for amplification as well as sequencing in one reaction (Berg and Olaisen, 1994)Degenerate PrimersDegeneracy in primer sequence should also be taken into consideration. In fact researchers pursuing the cloning of novel genes often face the problem that only a partial protein sequence is known (Bindon et al., 1998). In these circumstances several procedures can be used, some involveuniversal primers or reverse translation of the protein sequence into a DNA sequence and the design of primers from this sequence. However, due to redundancy in the genetic code, primer design must account for the ambiguous DNA bases and has to be designed in the region of lowest degeneracy (Kwok et al., 1994). Le Guyader et al. (1996) evaluated the effect of degenerate primers in the 生物秀-专心做生物w w w .b b i o o .c o mdetection of caliciviruses. Mack and Sninsky (1988) demonstrated the selection of conserved regions encoded by amino acids with minimal codon degeneracy in order to reduce mismatch. Degenerate primers based on the amino acid sequence of conserved regions were also used tosearch for members of a gene family (Wilks et al., 1989), homologous genes from different species (Kopin et al., 1990) and related viruses (Mack and Sninsky, 1988; Manos et al., 1989; Shih et al., 1989). A computer program was also developed specifically for degenerate primer design (Chen and Zhu, 1997).Software in Primer DesignMost molecular biological applications are aided by software. The use of software in biological applications has given a new dimension to the field of bioinformatics. Many different programs for the design of primers are now available. Freeware software is available on the internet and many universities have established servers where a user can log on and perform free analyses of proteins and nucleic acid sequences. There are number of simple stand-alone programs as well as complex integrated networked versions of the commercial software available. These software packages may be for complete DNA and protein analysis, secondary structure predictions, primer design, molecular modeling, development of cloning strategies, plasmid drawing or restriction enzyme analyses etc. Companies engaged in biosoftware development include: Alkami Biosystems, Molecular Biology Insights, PREMIER Biosoft International, IntelliGenetics Inc., Hitachi Inc., DNA Star, Advanced American Biotechnology and Imaging.Some scientists have also developed algorithms and computer programs for various purposes of primer design (Rychlik and Rhoades, 1989; Lowe et al., 1990; Lucas et al., 1991; O'Hara and Venezia, 1991; Tamura et al., 1991; Makarova et al., 1992; Osborne, 1992; Plasterer, 1997; Sze et al., 1998).ConclusionBiological science, and in particular biotechnology, is rapidly changing and cannot achieve its objectives without the help of computer technology and information technology tools. PCR primer design concepts are not new. However constant upgrading and updating of the strategies and methods are essential to maintain rapid and efficient progress. Computational strategies in biotechnology are of particular importance. The algorithms relevant to the efficient design of primers should be modified taking into account experimental data.生物秀-专心做生物w w w .b b i o o .c o mReferencesAlbert, J., and Fenyo, E.M. 1990. Simple, sensitive and specific detection of humanimmunodeficiency virus type 1 in clinical specimens by polymerase chain reactions with nested primers. J. Clin. Microbiol. 28: 1560-1564.Berg, E.S. and Olaisen, B. 1994. Hybrid PCR sequencing: sequencing of PCR products using a universal primer. Biotechniques 17: 896-901.Bindon, C., Martindale, J., and Mitchell, C. 1998. Biologically generated primer for PCR: PCR primer for unknown sequence. Nucleic Acid Res. 26(13): 3305-3358.Breslauer, K.J., Ronald, F., Blocker, H., and Marky, L.A.1986. Predicting DNA duplex stability from the base sequence. Proc. Natl. Acad. Sci. 83: 3746-3750.Bulat, S.A., Kobaev, O.K., Mironenko, N.V., Ibatullin, F.M., Luchkina, L.A., and Suslov, A.V. 1992. Polymerase chain reaction with universal primers for studying genomes. Genetika 28: 19-28. Chamberlain, J.S., Gibbs, R.A., Ranier, J.E., and Caskey, C.T. 1991. Detection of gene deletions using multiplex polymerase chain reactions, Meth. Molec. Biol. 9: 299-312.Chang, J.G., Lu, J.M., Huang, J., Chen, J.T., Liu, H.J., and Chang, C.P. 1995. Rapid diagnosis of b-thalassaemia by mutagenically separated polymerase chain reaction (MS-PCR) and its application to prenatal diagnosis, Br. J. Haemat. 91: 602-605.Chen, H., and Zhu,G.1997. Computer program for calculating the melting temperature of degenerate oligonucleotides used in PCR or hybridization. Biotechniques 21: 134-140Chou, Q., Russell, M., Birch, D.E., Raymond, J., and Bloch, W. 1992. Prevention of pre-PCRmis-priming and primer dimerization improves low-copy number amplifications. Nucleic Acids Res. 20: 1717-1723.Dieffenbach, C.W., Lowe, T.M.J., and Dveksler, G.S. 1993. General concepts for PCR primer design. In: PCR Methods and Applications, Cold Spring Harbor Laboratory 3: S30-S37.Drummond, R., and Gelfand, D.H. 1989. Isolation, characterization and expression in Escherichia coli of the DNA polymerase gene from Thermus aquaticus. J. Biol. Chem. 264: 6427-6436.Edwards, M.C., and Gibbs, R.A. 1994. Multiplex PCR: Advantages, development and applications. PCR Methods Applic. 3: S65-S75.Engelke, D.R., Hoener, P.A., and Collins, F.S. 1988. Direct sequencing of enzymatically amplified human genomic DNA. Proc. Natl. Acad. Sci. 85: 544-550.生物秀-专心做生物w w w .b b i o o .c o mErlich, H.A., Gefland, D., and Sninsky, J.J. 1991. Recent advances in the polymerase chain reaction. Science 252: 1643-1651.Freier, S.M., Kierzek, R., Jaeger, J.A., Sugimoto, N., Caruthers, M.H., Neilson, T., and Turner, D.H. 1986. Improved free-energy parameters for predictions of RNA duplex stability. Proc. Natl. Acad. Sci, USA. 83: 9373-9377.Green, C.E., Lund, J.K., and Manos, M.M. 1991. PCR amplification from parafin-embedded tissues: Recommendations on fixatives for long term storage and prospective studies. PCR Methods Appl. 1: 46-50.Haff La. 1994. Improved quantitative PCR using nested primers. PCR Methods Appl. 3: 322-327. He, Q., Marjamaki, M., Soini, H., Mertsola, J., and Viljanen, M.K. 1994. Primers are decisive for sensitivity of PCR. Biotechniques. 17: 82-87.Huang, M., Arnheim, N., and Goodman, M.F. 1992. Extension of base mispairs by Taq DNA polymerase: Implications for single nucleotide discrimination in PCR. Nucleic Acids Res. 20: 4567-4573.Hung, T., Mak, K. and Fong, K. 1990. A specificity enhancer for PCR, Nucleic Acids Res. 18: 4953-4959.Jung, V., Pestka, S.B., and Pestka, S. 1990. Efficient cloning of PCR generated DNA containing terminal restriction endonuclease sites. Nucleic Acid Res. 18: 6156-6160.Kain, K.C., Orlandi, P.A., and Lanar, D.E. 1991. Universal promoter for gene expression without cloning Expression-PCR. BioTechniques 10: 366-374.Kaneko, S., Feinstone, S.M., and Miller, R.H. 1989. Rapid and sensitive method for the detection of serum hepatitis B virus DNA using the polymerase chain reaction technique. J. Clin. Microbiol. 27: 1930-1933.Kawasaki, E.S. 1990a. Sample preparation from blood, cells and other fluids, In: PCR Protocols, a guide to methods and applications, Innis, M.A., Gefland, D.H., Sninsky, J.J. and White, T.J. (eds), Academic Press Inc., Berkeley CA, pp. 146-152.Kawasaki, E.S. 1990b. Amplification of RNA, In: PCR Protocols, a guide to methods andapplications, Innis, M.A., Gefland, D.H., Sninsky, J.J. and White, T.J. (eds), Academic Press Inc., Berkeley CA, pp. 21-27.Keller, G.H., Cumming, C.U., Huang, D.P., Manak, M.M., Ting, R. 1988. A chemical method for introducing Haptens onto DNA probes. Anal. Biochem. 170: 441-450.Klebe, R.J., Grant G.M., Grant, A.M., Garcia, M.A., Giambernardi, T.A., and Taylor, G.P. 1996. RT-PCR without RNA isolation. Biotechniques 21: 1094-1100.生物秀-专心做生物w w w .b b i o o .c o mKumar A., and Mishra B.N. 1997. Primer Premier 4. Biotech Software and Internet Journal 3: 31-38.Kwok S., Kellogg D.E., McKinney, E., Spasic, D., Levenson, C., Sninsky, J.J. 1990. Effects of Primer-template mismatches on the polymerase chain reaction: Human Immunodeficiency virus type I model studies, Nucleic Acids Res. 18: 999-1005.Kwok, S., Chang, S.Y., Sninsky, J.J., and Wong, A. 1994. A guide to the design and use of Mismatched and Degenerate primers. PCR Methods and Appl. S539-S547.Lawyer, F.C., Stoffel, S., Saiki, R.K., Myambo, K., Drummond, R., and Gelfand, D.H. 1989. Isolation, characterization, and expression in Escherichia coli of the DNA polymerase gene from Thermus aquaticus . J. Biol. Chem. 15: 6427-6437.Le Guyader, F., Estes, M.K., Hardy, E., Neill, F.H., Green, J., Brown, D.W., and Atmar, R.L. 1996. Evaluation of a degenerate primer for the PCR detection of human caliciviruses . Arch. Virol. 141: 2225-2235.Liang, P., and Pardee, A. 1992. Differential display of eukaryotic mRNAs by PCR. Science 257: 967-971.Linz, U., Delling, U., and Rubsamem-Waigmann 1990. Systematic studies on parametersinfluencing the performance of the polymerase chain reaction. J. Clin. Chem. Clin. Biochem. 28: 5-13.Liu, Q., Thorland E.C., and Sommer, S.S. 1997. Inhibition of PCR amplification by a point mutation down stream of a primer. Biotechniques 22: 292-298.Lowe, T., Shareifkin, J., Yang, S.Q., and Dieffenbach, C.W. 1990. A computer program for selection of oligonucleotide primers for polymerase chain reaction. Nucleic Acids Res. 18: 1757-1761.Lucas, K.M., Busch, S.M., and Thompson, J.A. 1991. An improved microcomputer program for finding gene or gene family-specific oligonucleotides suitable as primers for polymerase chain reactions or as probes. CABIOS 7: 525-529.Mack, D.H., and Sninsky, J.J. 1988. A sensitive method for the identification of uncharacterized viruses related to known virus groups: Hepadnavirus model system. Proc. Natl. Acad. Sci. 85: 6977-6981.Makarova, K.S., Mazin, A.V., Wolf, Y.I., and Soloviev, V.V. 1992. DIROM: An experimental design interactive system for directed mutagenesis and nucleic acid engineering. CABIOS 8: 425-431.生物秀-专心做生物w w w .b b i o o .c o m。

欧普德福 Mastercyclers产品说明书

Eppendorf Mastercyclers ®—best conditions for PCRAmplifyNew from Eppendorf:Mastercycler nexus X1Mastering PCR is easy—if you have the right equipment.Do not let poor instruments compromise your PCR results. Eppendorf blocks show outstanding homogeneity and accuracy. Fast ramp rates are precisely controlled allowing fast and reproducible PCR runs. Programming is easy and protocols can be password protected. All this supports your constant strive for reproducible and meaningful results.»Eppendorf—designinginstruments that fit the needs of today’s scientist.«Reproducibility > I n PCR, precise and accurate block control is paramount > T he Eppendorf blocks feature SteadySlope ® gradient technology to ensure that ramp rates are identical in both gradient and normal mode.Intuitivity > T he graphic programming is quick, easy and intuitive. > F ind all your protocols in your individual folder and protect them with a password. > P ause function, variable ramp rates and block modes and a lot more is at your disposal.Reliability > S pecially designed lids reduce the evaporation during PCR and help to accommodate a wide variety of PCR consumables. > T he solid handles allow one hand operation of the instrument. No turning knobs are needed to lower the heated lid.4Eppendorf Mastercycler FamilyMastercycler® nexus X1Learn more about the Mastercycler nexus X1 at /mastercycler5Eppendorf Mastercycler Family ArrayPure SilverThe new Mastercycler nexus X1 combines the modern and intuitive software from the Mastercycler nexus with a fast 96well silver block for increased heating and cooling rates. It is fast, easy to use, does not need much space or energy and sends you an email when it is done—what else do you need from a PCR cycler?Combine, Connect, Control>Combine up to 3 units for maximum throughput!>C onnect your Mastercycler nexus to your computer network and get a status email to your desk!>C ontrol all the relevant parameters of your PCR through the intuitive softwareDo you need a fast, intuitive and reliable PCR instrument? The Mastercycler nexus X1 is exactly that. It brings the reliability and ease of use of Mastercycler nexus at the speed of a silver block. Low energy consumption, easy expansion to 3 units and a small footprint make it even more attractive. Product features>Heating rate: 5°C/s>Fast silver block>Small footprint>Intuitive graphic programming>Up to two other units can be connected to a central unit>Email notification>fl exlid concept: automatic height adjustment of the lid allows you to use all types of consumables>2year warranty>Optional selftest functionalityApplications>Fast PCR>Standard PCR>Cycle sequencing6Eppendorf Mastercycler FamilyWhen you need even more flexibilitySpecial applications require special consumables. With its flat block and no wells, the Mastercycler nexus flat offers the optimal foundation for slides and other unconventional consumable formats.If you want to do in situ PCR, your results are normally influenced by the characteristic temperature transfer of the corresponding in situ adapter. The Mastercycler nexus flat can heat and cool your slides directly without the need for an adapter!Product features>Flat block without wells>Intuitive graphic programming >Small footprint>Up to two more units can be connected to the central unit >Email notification >2year warranty>Optional selftest functionality»To raise new questions, new possibilities, to regard old problems from a new angle, requires creative imagination and marks real advance in science.«Albert EinsteinCertified QualityAll Eppendorf cyclers follow these quality guidelines: >Individual, documented quality control certificates > C alibration accordingly to national and international standards: NIST (USA), DKD/PTB (Germany), UKAS/NPL (UK) >UL/cUL listedMastercycler ®nexus flat>For more information, visit /pcr7Eppendorf Mastercycler Family PCR next to meIn the age of networking, the new Mastercycler nexus is your reliable companion when it comes to the daily routine of PCR. It can accommodate 96well PCR plates, 0.2 mL PCR tubes, 0.2 mL PCR tube strips and 0.5 mL PCR tubes. It is easy to use, does not need much space or energy and sends you an email when it is done.Combine, Connect, Control>Combine up to 3 units for maximum throughput! > C onnect your Mastercycler nexus to your computer network and get a status email to your desk! > C ontrol all the relevant parameters of your PCR through the intuitive softwareIn good company—high quality consumablesEvery researcher doing PCR always wonders about the best instrument, the best master mix, the best polymerase. Also when it comes to choosing the plasticconsumables that build the connection between PCR instruments and your precious sample, the same rationality and prudence should be applied.Different consumables can make a huge difference in the quality and reproducibility of your PCR results. Wall thickness, thermal conductivity of the material, mechanical stability and many other technical characteristics will have a direct impact on your experiment and subsequently the results. Make sure you use the best PCR consumables for your application!>96well and 384well PCR plates for high and medium throughput >Divisible plates, PCR tubes and tube strips for lower throughput > S ealing options, racks and other accessories for an optimized workflowPCR ConsumablesMastercycler ®nexusProduct features>Universal block for plates, 0.2 mL and 0.5 mL PCR tubes >Small footprint>Intuitive graphic programming>Up to two other units can be connected to a central unit >Email notification>fl exlid ® concept: automatic height adjustment of the lid allows you to use all types of consumables >2year warranty>Optional selftest functionalityApplications >Standard PCR >Cycle sequencing>In situ PCR with adapterChoose the block format that is best for your PCR8Mastercycler® pro Eppendorf Mastercycler FamilyReproducibility at its bestGetting reproducible results quickly—that is vital for PCR in any application. The Mastercycler pro is unparalleled in its ability to fulfill this requirement. The vapo.protect™ concept reduces evaporation to a minimum. Therefore, concentrations in your PCR master mix remain consistent—minimizing nonspecific binding beyond importance. The high heating and cooling rates of the Mastercycler pro S give you the speed you need. Unsurpassed speed can be achieved with Impulse PCR, a hotstart function that increases heating rates to 8 °C/s.Stop evaporation effectively!The flexible fit of the fluid cushion minimizes evaporation of your reaction considerably. Only 0–3 % of the reaction volume evaporates using the new vapo.protect™technology.Thermocyclers of other suppliers show evaporation of up to 10 % in the center and up to >50 % at the corner positions of the block.Thus, the vapo.protect™ technology leads to improved reproducibility and specificity at the rim and in the corners of the block.9 Eppendorf Mastercycler FamilyUtmost FlexibilityThe Mastercycler pro can be run as a standalone unit. However, the Control Panel can control up to 5 different units, giving you higher throughput. If even higher throughput is needed, up to 30 units of any block format combination can be controlled with one computer. The advanced CycleManager pro software offers a client/server architecture, that enables one to check the status of the PCR from any computer in your local network. All collected data is archived in a database. If you are working in a GLP environment, your documentation needs will be strongly supported by the software.Product Features Mastercycler pro>Ultimate reduction of evaporation>Extremely fast heating and cooling rates>Intuitive graphic programming>Gradient blocks with SteadySlope technology>Up to 5 units can be networked using a Control Panel >Up to 30 units can be networked using a PC software >Display indicates cycler number in a network>2 year warranty>Optional selftest functionalityQuality is timeless1990Microcycler: Eppendorf introduces its fi rst thermal cycler using water to heat and cool.1997Mastercycler gradient: the fi rst gradient thermal cycler on the market.2003Mastercycler ep:Fast heating/cooling rates and ultra quiet operation—in a compact design.2005M astercycler ep realplex : Extremely fast optics for rapid data acquisition.2008Mastercycler pro:New vapo.protect ™technology reduces evaporation.20122013Mastercycler nexus Low noise, lowenergy consumption, email notifi cationMastercycler nexus X1Speed, low energyconsumption, low noise and email notifi cation1993M astercycler 5330:Eppendorf introduces the fi rst Mastercycler based on peltier technology.>to be continued …The Mastercycler gradient was Eppendorf’s fi rst gradient cycler. Today the gradient function remains critical for many PCR labs.The next generation Mastercycler pro is still one of the fastest cyclers around. Its vapo.protect™ concept is the latest in evaporation reduction technology.Eppendorf has now introduced the Mastercycler nexus and Mastercycler nexus X1. Email notifi cation, booking schedule, low noise levels and low energy consumption are only a few of the features ...Technical Specifications Mastercycler®proMastercycler®pro SMastercycler®pro 384Mastercycler®nexus gradientMastercycler®nexusMastercycler®nexus eco***Thermoblock Aluminum Silver Aluminum Aluminum Aluminum AluminumSample Capacity96 × 0.2 ml PCR tubesor 1 × 96well PCR plate 1 × 384wellPCR plate96 × 0.2 mL PCR tubesor 1 × 96well PCR plate or up to 71 x 0.5 mL PCR tubesTemperaturecontrol rangeof the block4–99 °C4–99 °CTemperaturecontrol modeFast, Standard, Safe Fast, Standard, SafeHeating technologyof the blockPeltier elements, Triple Circuit Technology Peltier elements, Triple Circuit Technology Gradient block over 12 columns over 24 columns over 12 columns––Gradient range1–20 °C1–24 °C1–20 °C1–20 °C––Gradienttemperature range30–99 °C30–99 °C––Lid temperaturerange37–110 °C37–110 °CLid descent and clos-ing pressure vapo.protect™ technology withThermal Sample Protectionflexlid® technology withThermal Sample ProtectionBlock homogeneity: 20 °C–72 °C95 °C ≤ ±0.3 °C≤ ±0.4 °C≤ ±0.3 °C≤ ±0.4 °CBlock temperatureaccuracy± 0.2 °C± 0.2 °CHeating rate*ca. 4 °C/s ca. 6 °C/s ca. 4 °C/s ca. 3 °C/sCooling rate*ca. 3 °C/s ca. 4,5 °C/s ca. 3 °C/s ca. 2 °C/sInterfaces Centronics, USB, CAN in, CAN out USB, Ethernet, CAN in, CAN out CAN in, CAN outDimensions(W × D × H)26 × 41.5 × 37 cm25 × 41.2 × 32.1 cmWeight18.5 kg (40.8 lbs)11 kg (24.2 lbs)10.5 kg (23.1 lbs) Power supply230 V, 50–60 Hz230 V, 50–60 HzMax. powerconsumption950 W700 WSound power levels≤ 56 dB(A)< 40 dB[A]* Heating and cooling rates measured at block**Unit can only be operated via a Mastercycler nexus unit (including flat, X1 versions) with control and display panelProduct appearance and/or specifications are subject to change without notice.Mastercycler®nexus gradienteco***Mastercycler®nexus flatMastercycler®nexus flat eco***Mastercycler®nexus GSX1Mastercycler®nexus SX1Mastercycler®nexus GSX1e***Mastercycler®nexus SX1e***Aluminum Aluminum Aluminum Silver Silver Silver Silver96 × 0.2 mL PCRtubes or 1 × 96well PCR plateor up to 71 x 0.5mL PCR tubes4 slides or equivalent96 × 0.2 ml PCR tubes or 1 × 96well PCR plate4–99°C4–99°CFast, Standard, Safe Fast, Standard, SafePeltier elements, Triple Circuit Technology Peltier elements, Triple Circuit Technologyover 12 columns––over 12 columns–over 12 columns–1–20 °C––1–20 °C–1–20 °C–30–99 °C––30–99 °C–30–99 °C–37–110 °C37–110 °Cflexlid® technology withThermal Sample Protectionflexlid® technology withThermal Sample Protection≤ ±0.3 °C≤ ±0.4 °C≤ ±0.3 °C≤ ±0.4 °C± 0.2 °C± 0.2 °Cca. 3 °C/s ca. 5 °C/sca. 2 °C/s ca. 3.5 °C/sCAN in, CAN out USB, Ethernet,CAN in, CAN outCAN in, CAN out USB, Ethernet, CAN in, CAN out CAN in, CAN out25 × 41.2 × 32.1 cm25 × 41.2 × 32.1 cm10.5 kg (23.1 lbs)11 kg (24.2 lbs)10.5 kg (23.1 lbs)11.2 kg (24.7 lbs)10.7 kg (23.6 lbs)230 V, 50–60 Hz230 V, 50–60 Hz700 W540 W≤ 40 dB(A)≤ 40 dB(A)G: Gradient blockS: Silver blocke: Eco version, needs to be connected to a Mastercycler nexus with control and display panelFrost & Sullivan European PCR Instrumentation Green Excellence Award 2012»Based on its recent analysis of the PCR instrumentation market, Frost & Sullivan recognizes Eppendorf with the 2012 European Green Excellence Award for its Mastercycler nexus. This ecofriendly instrument off ers exceptional powersaving capability, e.g. by a standby feature and the amount of power consumed overall.«Ordering informationInternational Order no. North America Order no.Mastercycler ® nexus with silver block Mastercycler ® nexus GSX16345 000.010 6345000028Mastercycler ® nexus SX16346 000.013 6346000021Mastercycler ® nexus GSX1e*6347 000.017 6347000025Mastercycler ® nexus SX1e*6348 000.010 6348000029Mastercycler ® nexus with universal block Mastercycler ® nexus gradient 6331 000.0176331000025Mastercycler ® nexus6333 000.0146333000022Mastercycler ® nexus gradient eco* 6334 000.018 6334000026Mastercycler ® nexus eco*6332 000.0106332000029Mastercycler ® nexus with flat block Mastercycler ® nexus flat6335 000.011 6335000020Mastercycler ® nexus flat eco*6330 000.013 6330000021Complete Mastercycler ® pro packages Mastercycler ® pro und Control Panel 6321 000.515950040015Mastercycler ® pro S und Control Panel 6325 000.510950040025Mastercycler ® pro 384 und Control Panel 6324 000.516950040035Individual Mastercycler ® pro modules Control Panel, incl. connection cable 6320 000.007950030050Mastercycler ® pro** 6321 000.019950030010Mastercycler ® pro S**6325 000.013950030020Mastercycler ® pro 384**6324 000.010*********AccessoriesCAN_BUS connection cable, 50 cm 5341 612.006 950014008CAN_BUS connection cable, 150 cm 5341 611.000 950014016Self test dongle6320 071.001950030040Temperature Verification System with 96 well sensor plate 6328 000.006 6328000006Temperature Verification System with 384 well sensor plate6328 000.0146328000014CycleManager pro, incl. installation manual, online help, and connection cable 5349 810.001 950017007CycleManager pro, incl. installation manual, online help5349 820.007950017202* To run a Mastercycler nexus with the suﬃ x »eco« or »e«, a Mastercycler nexus model without such a suﬃ x is needed. Up to 2 units with the suﬃ x »eco« or »e« can be connected to a Mastercycler nexus without such a suﬃ x. ** A Control Panel or CycleManager pro software (both sold separately) is required for operation. CAN_Bus connection cables are required to link cyclers together as a network.13Eppendorf Mastercycler Family/mastercyclerEppendorf , the Eppendorf logo, Eppendorf Mastercycler , flexlid , SteadySlope and Eppendorf twin.tec are registered trademarks of Eppendorf AG, Hamburg, Germany. vapo.protect™ is a trademark of Eppendorf AG, Hamburg, Germany. All rights reserved, including graphics and images. Copyright © 2012 by Eppendorf AG.Order No.: A533X12020/GB1/20T/1212/FEEL/STEFYour local distributor: /contact Eppendorf AG · 22331 Hamburg · Germany***********************·Ordering informationInternational Order no. North America Order no.0.1 mL Eppendorf PCR TubesPCR Tube Strips 0.1 mL, without caps (10 × 12 pieces)0030 124.8040030124804PCR Tube Strips 0.1 mL + Cap Strips, domed (10 × 12 pieces each) 0030 124.8120030124812PCR Tube Strips 0.1 mL + Cap Strips, flat (10 × 12 pieces each) 0030 124.82000301248200.2 mL Eppendorf PCR Tubes0.2 mL PCR Tubes, thinwalled with hinged lid (1000 tubes) 0030 124.3329510100068tube strip, for 0.2 mL PCR Tubes (10 × 12 pieces) 0030 124.3599510100220.5 mL Eppendorf PCR Tubes0.5 mL PCR Tubes, thinwalled with lid (500 tubes) 0030 124.5370030124537Cap Strips, Racks, Films & Foils Cap Strips, domed (10 × 12 pieces) 0030 124.8390030124839Cap Strips, flat (10 × 12 pieces) 0030 124.8470030124847PCR Rack, 10 pcs.0030 124.5450030124545Heat Sealing Film, 100 pcs. 0030 127.8380030127838Heat Sealing Foil, 100 pcs. 0030 127.8540030127854PCR Film (adhesive), 100 pcs.0030 127.7810030127781PCR Foil (adhesive), 100 pcs. 0030 127.7900030127790twin.tec PCR PlatesEppendorf twin.tec ® PCR Plate 96, skirted, clear 25 pcs.0030 128.648951020401Eppendorf twin.tec ® PCR Plate 96, semiskirted, clear 25 pcs.0030 128.575951020303Eppendorf twin.tec ® PCR Plate 96, unskirted low profile, clear 20 pcs.0030 133.3070030133307Eppendorf twin.tec ® PCR Plate 96, unskirted low profile, clear (divisible) 20 pcs. 0030 133.3580030133358Eppendorf twin.tec ® PCR Plate 96, unskirted (250 μL), clear, 20 pcs.0030 133.3660030133366Eppendorf twin.tec ® PCR Plate 96, unskirted (250 μL), clear (divisible) 20 pcs.0030 133.3740030133374Eppendorf twin.tec ® PCR Plate 384, clear, 25 pcs.0030 128.508951020702Eppendorf twin.tec ® microbiology PCR Plate 96, skirted, clear, 10 pcs.0030 129.3000030129300Eppendorf twin.tec ® microbiology PCR Plate 96, semiskirted, clear, 10 pcs. 0030 129.3260030129326Eppendorf twin.tec ® microbiology PCR Plate 384, clear, 10 pcs.0030 129.3420030129342For order numbers of additional twin.tec PCR Plates, please visit /pcr。

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以国际化的标准，致力于提供有关远洋全回转推进器相关问题的解决方案。

从公司的角度看来，Steerprop公司，将要成为全回转推进器项目技术的领导者。

Steerprop 全回转推进器的特点可靠性A great deal of effort has been put into Steerprop propulsors in order to ensure that they are the most dependable azimuth propulsors available. Reliability has an immense impact on the total lifecycle economy. The construction is kept as simple and fail-proof as possible.Steerprop公司为全回转推进器提供尽可能多的服务以保证他们的可靠性。

可靠性对于设备的耐用期限有很大的影响。

尽可能的简单且无故障的保存。

Flexibility has always been the basic philosophy behind the Steerprop propulsors. As a company, Steerprop Ltd. is able to provide owners and operators with exactly the products they require.灵活的操控模式一直是Steerprop公司全回转推进器的基础哲学。

作为一家公司，Steerprop公司有能力为船东精确地提供他们所需要的产品。

The vessel can turn around easily by using 360 all degrees propellers. They also benefit convenient control and the safety of navigation.采用360度全回转舵桨，船舶掉头灵活、原地回转、倒车快，操控方便，航行安全。

Propulsor efficiency and performance have an influence on both vessel performance (such as speed and bollard pull) and fuel consumption. Vessel performance and fuel consumption, in turn, have an impact on the total lifecycle economy.全回转的效率和性能对船的性能（例如速度和系柱拉力）以及耗油率都有影响。

船的性能以及耗油率，相应地，也会对总耐用期限产生影响。

360 all degrees azimuth proplusion is a kind of Z-drive configuration, which is energy-saving, of low noise and efficient device. With counter rotating propellers , the sailing speed can be raised , also the propulsion efficiency , meanwhile, owners could take advantage of the reducing fuel.360度全回转推进装置，是一种高效、节能、降噪、新颖的Z型船舶推进装置。

采用对转式推进器，可提高航速，提高推进效率，可节省燃料。

The propulsor with small stern vibration and average slipstream enables the vessel to sail in shallow waterways by using smaller diameter propellers. It also improve vacuole function .If using larger diameter propellers ,the propulsion efficiency would reduce.本推进器尾部振动减小，尾流流畅均匀，无横向推力，可用较小直径螺旋桨，适用航行于螺旋桨直接受到限制的浅水航道，并能改善空泡性能，克服螺旋桨直径较大和空泡影响推进效率的缺点。

• Power range 800...3500 kW; suited to the individual application and the rules of the selected classification society功率范围800-3500千瓦，适用于独立设备和指定船级社的规定• Open, ducted开放式的，导管式的• Z-drive or limited availability L-drive configurationsZ型传动方式或有限利用的L型传动方式• Diesel engine or electric motor prime mover柴油机或电动机作为原动机• Electric or hydraulic actuation of azimuth电动或液压驱动全回转• All major class society certificates available具有所有船级社的证书• Several mounting options: through hull from above, in two parts, or from below.多种安装方式：自上贯穿船体外壳，分两部分，或者从底下安装• Power range 4000...7000 kW; suited to the individual applications and the rules of the selected classification society功率范围4000-7000千瓦，适用于独立设备和指定船级社的规定• Open, ducted开放式的，导管式的• Z-drive configurationZ型传动方式• Electric motor prime mover电动机作为原动机• Electric actuation of azimuth电动驱动全回转• All major class society certificates available具有所有船级社的证书• Several mounting options: through hull from above, in two parts, or from below.多种安装方式：自上贯穿船体外壳，分两部分，或者从底下安装• Power range 800...4000 kW; suited to the individual application and the rules of the selected classification society功率范围800-4000千瓦，适用于独立设备和指定船级社的规定• Z-drive configurationZ型传动方式• Electric motor prime mover or direct diesel engine for low powers电动机作为原动机或低功率采用直流柴油机马达• Electric or hydraulic actuation of azimuth电动或液压驱动全回转• All major class society certificates available具有所有船级社的证书• Mounting options: in two parts or from below.安装方式：分两部分或者从底下安装• For ice going and arctic applications为扫冰和寒带地区使用的设备• Power range 800...16,000 kW; suited to the individual application and the rules of the selected classification society功率范围800-16,000千瓦，适用于独立设备和指定船级社的规定• Open pushing, ducted pushing, open pulling propellers, or CR Propellers开放式推动，管道式推动，开放推动式螺旋桨或者CR螺旋桨• Electric motor prime mover电动机作为原动机• Z-drive configurationZ型传动装置• For vessels that operate mostly in free-running conditions and also higher speeds.为那些大部分在自由且高速环境下运行的船只• Steerprop ECO propulsors with Contra-Rotating Propellers (CRP) offer the ideal solution, improving the fuel economy.Steerprop ECO 推进器和倒转推进器提供理想的解决方法，促进燃油的节省• The power capacity in this product category ranges from 5 to 20 MW这款产品类别范围内，功率范围为5-20兆瓦• The Steerprop ECO has a dual-end configuration which, combined with the sophisticated hydrodynamic design of the underwater parts, provides the highest efficiency in the propulsor industr y.Steerprop ECO有一个两端的配置，包括水下部分的精密的水动力设计，为推进器提供最高的推进效率The ability to control both the thrust and slipstream by a full 360 degrees makes the azimuth propulsor a powerful tool for ice management operations.控制推力和360度尾流的能力使得全回转推进器成为破冰的一个强大的工具。