ROX

LOL亚运会夺冠作文
亚运会在中国举办，作为全世界电竞爱好者共同期待的盛事，肯定少不了各种精彩绝伦的比赛。

昨天刚结束的英雄联盟比赛决赛可谓是百家争鸣、万众瞩目。

有亚洲最强赛区 LCK 的黑马 Rox 战队对抗世界上最大赛区的中国代表队 RNG。

虽然韩国战队一直以来都被视为实力超群，但面对 RNG 这样一支强劲的老牌劲旅还是让人捏把汗。

然而，作为 LPL 的头号种子选手， Uzi 最终带领队伍取胜。

可喜可贺！
第一局的比赛由韩国队拿下，首先由 Rox 的上单 Condi 拿出青钢影并且前期就打出优势，一波完美开团帮助 Rox 在一级就取得优势，接着更是强行配合中路秒掉 Knight。

而后利用峡谷先锋连推两塔拿到三条小龙，经济已遥遥领先。

最后 Rox 稳扎稳打凭借完美运营拿下比赛的胜利。

第二局比赛中， Rox 的中单 Jensen 再次祭出中单克烈强抓 Fly。

而反观 Fly 则是拿到自己擅长的佐伊去与对方中野周旋，双方爆发几波小规模团战，只见 Rune 残血开启加速冲进敌阵收割残血的画面简直令人瞠目结舌。

最后 Fly 以一个 E 闪收获四杀完成五杀拿下本场 MVP。

可能是我看得太专注了吧，从第二局比赛开始er 的英雄选择和操作就十分亮眼。

尤其是在生死局的最后一场，他的辛德拉又一次打出完美输出，操作了三次极限逃脱并将 Killua 控住随后连环爆炸，堪称技惊四座。

不知道是否由于这名选手年纪轻轻便斩获多项大奖而信心满满？所以第三局中他就掏出了杰斯这种传统 ADC。

本以为这位
韩服第一上单会如之前一般展现自己无解的“铁头功”，然而没想到他竟然在这种非常规情况下拿到一血。

6FAM-HEX-TMR-ROX四色荧光分析体系的构建刘莉;吴谨;侯一平;张霁;戴浩霖;廖淼;张林;陈国弟;李英碧【期刊名称】《中国法医学杂志》【年(卷),期】2006(021)005【摘要】目的为在310基因分析仪上进行6FAM-HEX-TMR-ROX四色荧光STR 复合扩增分型建立基础条件.方法以pGEM载体作为靶DNA,设计引物扩增获得500～80bp的13个长度不等DNA片段,以这些片段纯化物作为模板DNA,分别扩增获得6FAM、HEX、TMR和ROX标记的Matrix标准物,用4种Matrix标准物在310基因分析仪上构建可用于6FAM-HEX-TMR-ROX四色荧光分析的Matrix 文件.结果扩增所得的13个非标记片段,长度分别为80、100、120、140、160、180、200、220、260、300、340、400、500bp,在非变性聚丙烯酰胺凝胶连续电泳-银染凝胶上均表现为单条带,6FAM、HEX、TMR、ROX分别标记的片段通过310基因分析仪检测均为单峰.混合ROX标记的80、120、180、220、260、300bp的6个片段获得的内标,显示出良好的线性关系.结论建立了有效的6FAM-HEX-TMR-ROX四色荧光分析Matrix,为进行多个STR基因座荧光标记复合扩增检测奠定了基础.【总页数】5页(P274-277,彩插Ⅱ)【作者】刘莉;吴谨;侯一平;张霁;戴浩霖;廖淼;张林;陈国弟;李英碧【作者单位】四川大学华西基础医学与法医学院,四川,成都,610041;四川大学华西基础医学与法医学院,四川,成都,610041;四川大学华西基础医学与法医学院,四川,成都,610041;四川大学华西基础医学与法医学院,四川,成都,610041;四川大学华西基础医学与法医学院,四川,成都,610041;四川大学华西基础医学与法医学院,四川,成都,610041;四川大学华西基础医学与法医学院,四川,成都,610041;四川大学华西基础医学与法医学院,四川,成都,610041;四川大学华西基础医学与法医学院,四川,成都,610041【正文语种】中文【中图分类】D9【相关文献】1.3个miniSTR基因座四色荧光复合分型体系的构建及应用 [J], 刘宁;左敏;李淑瑾;丛斌;白雪;谷建立;侯志平2.四色荧光DNA测序中的转换矩阵分析 [J], 林云跃;严惠民;王立强3.河北汉族人群D9S1122/D10S1435/D12ATA63 miniSTR基因座四色荧光标记分型体系的构建及遗传多态性研究 [J], 侯志平;丛斌;李淑瑾;白雪;谷建立;刘宁4.树突状细胞前体细胞亚群四色荧光分析方法探讨 [J], 李玉珠;史敦云;张琼丽5.毛细管电泳四色荧光检测法分析茶树SSR标记 [J], 黄建安;李娟;谭月萍;刘硕谦;李勤;刘仲华因版权原因，仅展示原文概要，查看原文内容请购买。

I. Introduction5X PrimePath™ Probe qPCR Kit, ROX plus, GPR (Cat. Nos. 638348 & 638350) enables the user to perform accurate qPCR on DNA samples. The highly concentrated master mix allows for flexibility in handling largersample volumes. The entire protocol from setup to detection takes less than one hour, enabling fast results. II. Required MaterialsThis protocol applies to the following Takara Bio products:•5X PrimePath Probe qPCR Kit, ROX plus, GPR (Cat. Nos. 638348 & 638350)Additional materials required:•Primers and probes•Micropipette and tips (with hydrophobic filters)•Vortex mixer•Benchtop centrifuge for tubes or plates• 1.5 ml Eppendorf tubes, 200 μl PCR tubes, or 200 μl PCR plates for sample preparation•Tubes or plates for real-time PCR• A real-time PCR machine compatible with low ROX (e.g., QuantStudio 3 or later, ABI 7500, ABI 7500 Fast, ViiA 7, Stratagene MX4000P, MX3000P, MX3005P, etc.)NOTE: If using a high ROX qPCR instrument, add ROX based on the guidelines for your model and theconcentration of the ROX additive used.III. Protocol1.Mix the tube of 5X PrimePath Probe qPCR Mix by inverting several times then spin down for 10 sec; the mixmay appear cloudy after storage, but this does not affect its performance.2.On ice and using the table below, combine everything for the qPCR reaction mix EXCEPT the DNA samplefor all planned reactions, plus 10% of the total reaction mix volume for overage.NOTES:–If the mixture is used immediately, it can be combined at room temperature (no ice).–Do not add the sample at this time. It is listed in the table to indicate what the final mixture will contain.qPCR reaction mix (per 25 µl* reaction)Reagent Finalconcentration Singleplex Multiplex(N† targets)5X PrimePath Probe qPCR Mix, ROX plus, GPR 1X 5 µl 5 µl PCR Forward Primer (10 μM)0.2 μM0.5 µl N† x 0.5 µl PCR Reverse Primer (10 μM)0.2 μM0.5 µl N† x 0.5 µl Probe (10 μM)0.2 μM0.5 µl N† x 0.5 µl DNA sample‡— 2 µl§ 2 µl§RNase Free H2O — Up to 25 µl**Up to 25 µl** Total volume per reaction25 µl25 µl*Mix, as written, can also be used per 20 µl reaction.†Where N represents the total number of targets of interest. The total reaction volume will still be 25 µl, while 0.5 µl of each primer and probe will be added per target.‡Do not add to the reaction mix; this will be added to the mix in Step 4.§The most common sample volume of Takara Bio PCR products. It is possible to add more; however, depending on the chemicals contained in the sample solution, higher sample volume may have adverse effects.**Adjust the reaction volume according to the recommendations for the real-time PCR instrument used.3.Add [25 μl – Volume of DNA Sample] of the qPCR reaction mixture into a PCR tube or a 96-well PCR plate.4.Dispense the volume of DNA sample into the PCR tube or plate well. Spin down for 10 sec.5.Set up the thermal cycler and run the assay using the reaction conditions below:95°C 20 sec40 cycles:95°C 1 sec60°C 20 secNOTE: Please follow the instruction manual of the real-time qPCR machine used. If the default setting doesnot work, perform manual analysis per the instruction manual.6.After the reaction is complete, check the amplification curve. Confirm that the analytical parameters arecorrect and that the Ct value has been calculated.Contact UsCustomer Service/Ordering Technical Supporttel: 800.662.2566 (toll-free) tel: 800.662.2566 (toll-free)fax: 800.424.1350 (toll-free) fax: 800.424.1350 (toll-free)web: /service web: /supporte-mail: **********************e-mail: *******************************Notice to PurchaserThis product is a general purpose reagent intended For Laboratory Use. Outside of the United States, this product is intended for research use only unless otherwise stated. This product is not intended for a specific application or made for any particular clinical use. The performance characteristics of this product have not been fully established. It is the user´s responsibility to validate the performance of the product, and any component thereof, for any particular use. Resale or transfer of this product, any component thereof, or any substance produced through use of this product, or any component thereof, to any third party is expressly forbidden. To obtain additional rights, please contact your local office or go to our website for additional information.Your use of this product is also subject to compliance with the licensing requirements, listed below if applicable, and described on the product´s web page at. It is your responsibility to review, understand and adhere to any restrictions imposed by these statements.© 2023 Takara Bio Inc. All Rights Reserved.All trademarks are the property of Takara Bio Inc. or its affiliate(s) in the U.S. and/or other countries or their respective owners. Certain trademarks may not be registered in all jurisdictions. Additional product, intellectual property, and restricted use information is available at .This document has been reviewed and approved by the Quality Department.。

固相萃取-超高效液相色谱三重四级杆质谱联用法测定水中四环素、土霉素及罗红霉素尹燕敏【摘要】A rapid analytical method for simultaneous determination of tetracyclin, oxytetracycline and roxithromycin in water sample was developed. Water samples were purified and concentrated by solid phase extraction(SPE), then analyzed by ultra performance liquid chromatography electrospray tandem mass spectrometry(UPLC-MS/MS). Using formic acid and acetonitrile as the mobile phase, the three antibiotics were separated within 5 min. The limit of determination for anlytes were 0.08-0.35 ng/L with the relative standard deviation of 1.4%-5.6%. The recoveries of the antibiotics were in the range of 82.5%-114%for blank and 71.5%-126%for real samples.%建立了固相萃取-超高效液相色谱三重四级杆质谱联用法同时测定水中的罗红霉素、四环素和土霉素残留。

水样经过固相萃取纯化、富集，液质联用分析，采用甲酸溶液和乙腈作为流动相，在5 min 内完成对3种目标化合物的分析，3种目标化合物的方法检出限介于0.08～0.35 ng/L之间，测定结果的相对标准偏差为1.4%~5.6%，空白样品和实际样品的加标回收率分别为82.5%～114%，71.5%～126%。

QPCR FAQ（四）- 常见的曲线异常2017-09-08 16:02诺唯赞原标题：QPCR FAQ（四）- 常见的曲线异常小V是真心地希望并祝福科研宝宝们实验都顺顺利利~可科研做多了，总会遇到各种奇葩结果，比如下面这些情况……不用担心，小V帮你找原因。

1、扩增曲线形状异常a、扩增曲线不光滑：信号太弱，经系统矫正后产生。

提高模板浓度重复实验。

、扩增曲线断裂或下滑：模板浓度较高，基线的终点值大于Ct值。

减小基线终点 (Ct值-4)，重新分析数据。

图1 基线设置不当扩增曲线c、个别扩增曲线突然骤降：反应管内留有气泡，由于温度升高后气泡破裂，使仪器检测到的荧光值突然降低所致。

进行扩增反应之前要仔细检查反应管内是否有气泡残留。

d、扩增曲线呈锯齿状且不连续：ROX添加不当，需校正参比染料。

2、反应结束无扩增曲线出现a、反应循环数不够：一般设置循环数为40，但需要注意的是过多的循环会增加过多的背景信号，降低数据可信度。

、确认程序中是否设置了信号采集步骤：两步法扩增程序一般将信号采集设置在退火延伸阶段；三步法扩增程序应当将信号采集设置在72°延伸阶段（如蓝色箭头所示）。

c、确认引物是否降解：长时间未用的引物应先通过PAGE电泳检测完整性，以排除引物降解的可能性。

d、模板浓度太低：减少稀释度重复实验，一般未知浓度的样品先从最高浓度做起。

e、模板降解：重新制备模板，重复实验。

3、溶解曲线形状异常a、引物设计不佳①重新设计引物。

②若杂峰为引物二聚体，可尝试提高模板浓度、退火温度或者降低引物浓度进行改善。

③当目的基因表达量极低时，染料法QPCR容易形成引物二聚体产物影响实验结果，此时，建议使用探针法定量。

、ROX使用不当图9 ROX添加不当溶解曲线图怎样避免ROX使用不当？①看清标签以防高、低浓度ROX拿错。

②待ROX完全解冻且混匀后吸取添加。

如果您有遇到更奇葩的异常曲线存在疑惑，可以直接拨打诺唯赞技术支持热线进行咨询哦~（更多详细内容敬请留意Vazyme 针对QPCR的讲座信息。

07/08第二学期药物合成反应试题一、填空题（0.5分×40＝20分）1、轨道的节越多，它的能量越 高 。

2、1S 轨道呈 球 对称，对应于氢原子的电子的 最低可能的轨道 。

3、写出以下原子的基态电子结构：Be: 1S22S2 C: 1S22S22Px12Py1 F: 1S22S2SPx22Py22Pz1 Mg: 1S22S22Px22Py22Pz23S2 4、据分子轨道理论，两个原子轨道重叠形成两个分子轨道，其中一个叫成键轨道 ，具有比原来的原子轨道更低的能量；一个叫反键轨道 ，具有较高的能量。

5、场效应和分子的几何构型 有关，而诱导效应只依赖于键 。

6、碳－碳键长随S 成分的增大而变短 ，键的强度随S 成分的增大而增大 。

7、判断诱导效应（－Is ）强弱：－F,－I,－Br,－Cl 。

－F>－Cl>－Br>－I 8、判断诱导效应（＋Is ）强弱：－C(CH 3)3>－CH(CH 3)2>－CH 2CH 3>－CH3。

9、一般情况下重键的不饱和程度愈大，吸电性 愈强 ，即－Is 诱导效应愈大。

10、在共轭体系中，形成共轭体系的原子以及和这些原子直接结合的原子都在同一平面里 ；各个键上的电子云密度在一定程度上都趋于平均化 。

11、布朗斯特酸的定义是给出质子的物质 ，路易斯酸的定义是有空轨道的任何一种物质 。

12、卤素对双键亲电加成的活性顺序是：Cl 2>Br 2>I 2 。

13、卤素对双键亲电加成是按分步机理进行的， X +先对双键做亲电进攻，而后X _再加上去。

14、判断共轭类型，并用箭头标明电子云偏移方向：π－π共轭p －π 共轭p －π 共轭15、溴素或氯对烯烃的加成一般属于 亲电加成 机理。

16、在卤素对烯烃的加成反应中，当双键上有苯基取代时，同向加成的机会 增C(CH 3)3CH 2CH 3CH 3CH(CH 3)2C H CH 2CH 2H 2CH加 。

©2018 QIAGEN Beverly, Inc. 100 Cummings Center Suite 407J Beverly, MA 01915 Quantabio brand products are manufactured by QIAGEN Beverly, Inc.Intended for molecular biology applications. This product is not intended for the diagnosis, prevention or treatment of a disease.95139 / IFU-095.1 REV 02PerfeC T a ® qPCR ToughMix ® UNG ROX ™DescriptionPerfeC T a qPCR ToughMix UNG ROX is a 2X concentrated ready-to-use reaction cocktail for PCR amplification of DNA templates that overcomes many known inhibitors of PCR often present in crude samples extracted from environmental specimens, plant tissues, or animal tissues. It is a versatile and robust real-time qPCR reagent that provides maximum sensitivity and PCR efficiency with a variety of fluorogenic probe chemistries, including TaqMan ® hydrolysis probes. PerfeC T a qPCR ToughMix UNG ROX contains all required reaction components, except primers, probe(s), and DNA template. Inclusion of uracil DNA glycosylase (UNG), and substitution of dTTP with dUTP, prevents amplification of carry-over contamination from previous dU-containing PCRs.A key component of PerfeC T a qPCR ToughMix UNG ROX is an ultra pure, highly processive thermostable DNA polymerase that is combined with high avidity monoclonal antibodies. This proprietary polymerase mix is highly resistant to PCR inhibitors and provides an extremely stringent automatic hot-start allowing reaction assembly, and temporary storage, at room temperature prior to PCR amplification. PerfeC T a qPCR ToughMix UNG ROX delivers exceptional performance with either fast or conventional PCR cycling protocols.Instrument CompatibilityDifferent real-time PCR systems employ different strategies for the normalization of fluorescent signals and correction of well-to-well optical variations. It is important to match the appropriate reference dye to each specific optical detection system. PerfeCta qPCR ToughMix, ROX contains an optimal concentration of a stabilized carboxy-X-rhodamine compound (ROX ™) for instruments that use an excitation wavelength of ~490 nm and 605 to 610 nm emission channel for the reference signal. Please consult our Product Finder selection tool at to find the correct product for your real-time PCR system. ComponentsPerfeC T a qPCR ToughMix UNG ROX (2X): 2X concentrated reaction buffer containing optimized concentrations of MgCl 2, dNTPs (dATP, dCTP, dGTP,dUTP), hot-start DNA polymerase, uracil DNA glycosylase (UNG), and stabilizers.Storage and StabilityStore components in a constant temperature freezer at -25°C to -15°C protected from light upon receipt. Repeated freezing and thawing does not impair product performance. After thawing, mix thoroughly before using.For lot specific expiry date, refer to package label, Certificate of Analysis or Product Specification Form.Guidelines for qPCR:▪ The design of highly specific primers and probes is a critical parameter for successful real-time PCR. The use of computer aided primer design programs is encouraged in order to minimize the potential for internal secondary stru cture and complementation at 3’-ends within each primer, the primer pair, and primer/probe combinations. For best results, amplicon size should be limited to 65 - 200 bp. Optimal results may require titration of primer concentration between 100 and 900 nM. A final concentration of 300 - 400 nM each primer and 100 to 250 nM probe is effective for most applications. Increasing the concentration of the primer that initiates synthesis of the target strand that is complementary to the probe can improve fluorescent signal for some primer/probe systems.▪ Preparation of a reaction cocktail is recommended to reduce pipetting errors and maximize assay precision. Assemble the reaction cocktail with all required components except sample template (genomic DNA or cDNA) and dispense equal aliquots into each reaction tube. Add the DNA template to each reaction as the final step. Addition of samples as 2 to 5- L volumes will improve assay precision.▪ Suggested input quantities of template are: cDNA corresponding to 1 pg to 100 ng of total RNA; 10 pg to 1 µg genomic DNA▪ After sealing each reaction, vortex gently to mix contents. Centrifuge briefly to collect components at the bottom of the reaction tube. Reaction AssemblyComponentVolume for 20-μL rxn.Final ConcentrationPerfeC T a qPCR ToughMix UNG ROX (2X) 10 µL 1xForward primer variable 100 – 900 nM Reverse primer variable 100 – 900 nM Probevariable 100 – 250 nMNuclease-free water variable Template2 – 5 µL variable Final Volume (μL)20 µLNote : For smaller or larger reaction volumes scale all components proportionally.Cat No. 95139-250 Size: 250 x 20-uL reactions (2 x 1.25 mL) Store at -25ºC to - 15°C protected from light95139-012 1250 x 20-µL reactions (10 x 1.25 mL)95139-05K5000 x 20-µL reactions (1 x 50 mL)©2018 QIAGEN Beverly, Inc. 100 Cummings Center Suite 407J Beverly, MA 01915 Quantabio brand products are manufactured by QIAGEN Beverly, Inc.Intended for molecular biology applications. This product is not intended for the diagnosis, prevention or treatment of a disease.95139 / IFU-095.1 REV 02PCR Cycling ProtocolFast 2-Step Cycling Fast 3-Step Cycling Standard Cycling UNG carry-over incubation – optional:Initial denaturation: PCR cycling (30-45 cycles):The appropriate step for fluorescent data collection varies for different probe assay formats. Data collection for 5’-nuclease probe assays (TaqMan probe) should be carried out at the end of the extension step. Use the annealing step for data collection with hybridization probe assays (HybProbe ® FRET hybridization probes, Molecular Beacons, Solaris ® qPCR Assays, Scorpions ® primers, etc.).End-point analysis should be carried out at a suitable temperature for your detection probe chemistry.‡ UNG incubation is optional. Alternate protocols are acceptable. We find that a 5 minute incubation at 45ºC is significantly more effective at eliminating carry-over contamination than the more typical procedure of 50ºC for 2 min.* Full activation of the DNA polymerase occurs within 10 seconds at 95ºC; however, optimal initial denaturation time is template dependent and will affect qPCR efficiency and sensitivity. Amplification of genomic DNA or supercoiled plasmid DNA targets may require 5 to 10 min at 95ºC to fully denature and fragment the template. Short double-stranded DNA template (PCR product) or single-stranded DNA template, such as cDNA, may require as little as 1s at 95ºC. Use 30s at 95ºC as a general starting point.† Extension time is dependent upon amplicon length and the minimal data collection time requirement for your qPCR instrument. Use 30s at 60ºC as a general starting point. Some assay designs and/or detection chemistries may require a 3-step cycling protocol for optimal performance. Optimal annealing temperature and time may need to be empirically determined for any given primer set and real-time instrument.Quality ControlKit components are free of contaminating DNase and RNase. PerfeC T a qPCR ToughMix UNG ROX is functionally tested in qPCR. Kinetic analysis must demonstrate linear resolution over six orders of dynamic range (R 2 > 0.990) with a 2-fold discrimination of starting template and a PCR efficiency > 95%.Limited Label LicensesUse of this product signifies the agreement of any purchaser or user of the product to the following terms:1. The product may be used solely in accordance with the protocols provided with the product and this manual and for use with components contained in the kit only. QIAGEN Beverly, Inc. grants no license under any of its intellectual property to use or incorporated the enclosed components of this kit with any components not included within this kit except as described in the protocols provided with the product, this manual, and additional protocols available at Some of these additional protocols have been provided by Quantabio Product users for Quantabio users. These protocols have not been thoroughly tested or optimized by QIAGEN Beverly, Inc.. QIAGEN Beverly, Inc. neither guarantees them nor warrants that they do not infringe the rights of third-parties.2. Other than expressly stated licenses, QIAGEN Beverly, Inc. makes no warranty that this kit and/or its use(s) do not infringe the rights of third-parties.3. This kit and its components are licensed for one-time use and may not be reused, refurbished, or resold.4. QIAGEN Beverly, Inc. Specifically disclaims any other licenses, expressed or implied other than those expressly stated.5. The purchaser and user of the kit agree not to take or permit anyone else to take any steps that could lead to or facilitate any acts prohibited above. QIAGEN Beverly, Inc. may enforce the prohibitions of this Limited License Agreement in any Court, and shall recover all its investigative and Court costs, including attorney fees, in any action to enforce this Limited License Agreement or any of its intellectual property rights relating to the kit and/or its components.This product is licensed under U.S. Patent No. 7,972,828 and corresponding US and foreign patents and patent applications for any use for research and development purposes; the license expressly excludes any use for diagnostic testing or clinical therapeutics in humans or animals.The use of this product is covered by at least one claim of U.S. Patent No. 7,687,247 owned by Life Technologies Corporation. The purchase of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product in research conducted by the buyer (whether the buyer is an academic or for-profit entity). The buyer cannot sell or otherwise transfer (a) this product, (b) its components, or (c) materials made by the employment of this product or its components to a third party or otherwise use this product or its components or materials made by the employment of this product or its components for Commercial Purposes. Commercial Purposes means any activity for which a party receives or is due to receive consideration and may include, but is not limited to: (1) use of the product or its components in manufacturing; (2) use of the product or its components to provide a service, information, or data; (3) use of the product or its components for therapeutic, diagnostic or prophylactic purposes; or (4) resale of the product or its components, whether or not such product or its components are resold for use in research. The buyer cannot use this product or its components or materials made using this product or its components for therapeutic, diagnostic or prophylactic purposes. Further information on purchasing licenses under the above patents may be obtained by contacting the Licensing Department, Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008. Email: *************************.PerfeC T a, and ToughMix are registered trademarks of QIAGEN Beverly, Inc.. TaqMan is a registered trademark of Roche Molecular Systems, Inc. HybProbe is a registered trademark of Roche Diagnostics GmbH. ROX is a trademark Life Technologies Corporation. Solaris is a registered trademark of Thermo Fisher Scientific Inc. Scorpions is a registered trademark of DxS, Ltd. of Manchester, UK.。

河南农业2021年第25期ZHILIANG ANQUAN质量安全1.生物安全柜内消毒。

如进行“非洲猪瘟”检测活动时应在生物安全柜中操作区铺隔水垫，实验后卷起隔水垫置于污物桶后消毒灭菌，采用有效消毒剂将生物安全柜内操作区消毒处理1遍，再用清水浸泡纱布或吸水纸清洗1~2 遍。

2.剪刀、镊子等可重复使用物品消毒。

剪刀、镊子等可高压灭菌的物品直接置于锐器盒高压灭菌处理。

不能高压灭菌的物品考虑消毒剂浸泡，可用塑料盒盛装有效消毒剂进行浸泡处理 30 min。

不能用消毒剂浸泡但可高压处理的，可放进生物安全灭菌袋后直接高压蒸汽消毒（121 ℃，30 min）。

3.仪器、工作台面、地面、环境消毒。

消毒的频次可根据做实验次数的多少来定，少时14 d 消毒1次，多时7 d 消毒1次。

可用紫外线灯消毒30~60 min，但紫外线灯用久后不能起到杀灭微生物的效果，所以实30 说明书，检查荧光来作为参比荧光染ROX 来作为参比荧批荧光反应管内的体积误差等。

目前，市面上有的荧光定量PCR 试剂盒预混液是不含有ROX 参比荧光染料的，如果用此类试剂盒时，应将ROX 参比功能关闭，否则可能会出现围绕基线上下波动的“锯齿状”扩增曲线。

遇到这种问题，可以在“菜单设置”中找到“参比荧光染料”功能，将ROX 改为None，重新将样本加入含荧光反应液和聚合酶的反应体系中重新进行扩增分析，结果就正常了。

切记不要将原来的反应管重新进行扩增1次，如果这样，扩增曲线是一条直线，结果呈阴性，从而造成检验结果的错误。

有的荧光定量PCR 仪默认是对所有荧光通道及所有样品孔进行荧光采集，用户不用害怕数据丢失，实验结束后，可对仪器设置错误的反应孔的荧光通道、样品名称等进行更改，结果就会显示出正确的扩增曲线。

还有可能出现其他一些异常结果。

如多组分图扩 淮滨县动物疫病预防控制中心 周爱军 淮滨县畜产品检验检测中心 程泽影河南农业2021年第25期ZHILIANG ANQUAN质量安全增曲线没有抬升，本应是很明显的阴性样本，但是扩增图谱的扩增曲线抬升了，这样抬升曲线就会与阈值线相交，反而出现了CT 值。

Catalog No. RT0411-01产品简介2×SYBR Green Mix（With ROX）是使用染料法（SYBR GreenⅠ）进行实时荧光定量PCR扩增反应的预混体系，包括Hotstart Taq DNA Polymerase，Buffer，dNTPs，SYBR Green I荧光染料、Mg2+和ROX校正染料。

本品含有经化学修饰的全新高效的热启动酶Hotstart Taq DNA Polymerase，在常温下没有聚合酶活性，有效避免在常温条件下由引物和模板非特异性结合或引物二聚体而产生的非特异性扩增，酶的激活须在95℃下孵育10 分钟。

主要用于基因组DNA靶序列和RNA反转录后cDNA靶序列的检测。

本品所含的荧光染料SYBR Green I可以与所有的双链DNA结合，使该产品可用于不同靶序列的检测而不需合成特异性标记探针。

所含的ROX染料可校正定量PCR仪孔与孔之间产生的荧光信号误差, 适用于以ROX作为校正染料的所有荧光定量PCR仪。

产品包装保存条件-20℃避光保存，尽量避免尽量避免反复冻融。

注意事项1 使用前请上下颠倒轻轻混匀，尽量避免起泡，并经短暂离心后使用。

2 本产品中含有SYBR Green I 荧光染料和ROX 染料，保存本产品或配制PCR 反应液时应避免强光照射。

3 避免反复冻融本品，反复冻融可能使产品性能下降。

如果在短期内需要频繁使用，可在2-8℃保存。

使用方法以下举例为常规PCR 反应体系和反应条件，实际操作中应根据模板、引物结构和目的片段大小不同进行相应的改进和优化。

1.PCR反应体系：注：引物浓度以0.1-1.0 μM终浓度作为设定范围的参考。

扩增效率不高时，提高引物的浓度；发生非特异性反应时，可降低引物浓度; DNA模板的量以10-100 ng基因组DNA或1-10 ng cDNA为参照。

2. PCR反应程序：两步法PCR：步骤温度时间预变性95℃10 min变性95℃15 s退火/延伸60℃ 1 min融解曲线分析95℃15 s60℃ 1 min95℃15 s60℃15 s注意：1）本产品所采用的热启动酶须在预变性95℃、10 min条件下实现酶的活化。