Name: dna polymerase activity assay using near-infrared fluorescent labeled dna visualized by acrylamide gel electrophoresis
Uploaded: 2017-10-06T15:00:00.000+00:00
Duration: 7 min 38 s
Description: Watch this Scientific Journal Video about DNA Polymerase Activity Assay Using Near-infrared Fluorescent Labeled DNA Visualized by Acrylamide Gel Electrophoresis at JoVE.com

这项工作得到了科学进步研究公司 (科特雷尔学院学者奖 #22548) 和三生物技术 (ResearchReward 赠款 #G139) 的支持。

<ol>
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L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design+and+discovery+of+new+combinations+of+mutant+DNA+polymerases+and+modified+DNA+substrates.">Design and discovery of new combinations of mutant DNA polymerases and modified DNA substrates.</a> Chembiochem. 18 (8), 816-823 (2017).</li><li>Creighton, S., Goodman, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gel+kinetic+analysis+of+DNA+polymerase+fidelity+in+the+presence+of+proofreading+using+bacteriophage+T4+DNA+polymerase.">Gel kinetic analysis of DNA polymerase fidelity in the presence of proofreading using bacteriophage T4 DNA polymerase.</a> J Biol Chem. 270 (9), 4759-4774 (1995).</li><li>Joyce, C. M., Benkovic, S. 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R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=DNA+sequencing+with+chain-terminating+inhibitors.">DNA sequencing with chain-terminating inhibitors.</a> Proc Natl Acad Sci U S A. 74 (12), 5463-5467 (1977).</li><li>Karger, B. L., Guttman, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=DNA+Sequencing+by+Capillary+Electrophoresis.">DNA Sequencing by Capillary Electrophoresis.</a> Electrophoresis. 30 (Suppl 1), S196-S202 (2009).</li><li>Lawyer, F. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-level+expression,+purification,+and+enzymatic+characterization+of+full-length+Thermus+aquaticus+DNA+polymerase+and+a+truncated+form+deficient+in+5'+to+3'+exonuclease+activity.">High-level expression, purification, and enzymatic characterization of full-length Thermus aquaticus DNA polymerase and a truncated form deficient in 5' to 3' exonuclease activity.</a> PCR Methods Appl. 2 (4), 275-287 (1993).</li><li>Lawyer, F. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation,+characterization,+and+expression+in+Escherichia+coli+of+the+DNA+polymerase+gene+from+Thermus+aquaticus.">Isolation, characterization, and expression in Escherichia coli of the DNA polymerase gene from Thermus aquaticus.</a> J Biol Chem. 264 (11), 6427-6437 (1989).</li><li>Carroll, S. S., Cowart, M., Benkovic, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+mutant+of+DNA+polymerase+I+(Klenow+fragment)+with+reduced+fidelity.">A mutant of DNA polymerase I (Klenow fragment) with reduced fidelity.</a> Biochemistry. 30 (3), 804-813 (1991).</li><li>Shendure, J., Ji, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Next-generation+DNA+sequencing.">Next-generation DNA sequencing.</a> Nat Biotechnol. 26 (10), 1135-1145 (2008).</li><li>Larsen, A. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+general+strategy+for+expanding+polymerase+function+by+droplet+microfluidics.">A general strategy for expanding polymerase function by droplet microfluidics.</a> Nat Commun. 7, 11235 (2016).</li><li>Cozens, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enzymatic+Synthesis+of+Nucleic+Acids+with+Defined+Regioisomeric+2'-5'+Linkages.">Enzymatic Synthesis of Nucleic Acids with Defined Regioisomeric 2'-5' Linkages.</a> Angew Chem Int Ed Engl. 54 (51), 15570-15573 (2015).</li></ol>

作者没有什么可透露的。

在这里, 我们描述了一种测定 dna 聚合酶介导的 M dna 的合成。利用近红外标记的 DNA 引物, 利用变性聚丙烯酰胺凝胶电泳解决不同大小的寡核苷酸, 可以获得核苷酸的单核苷酸分辨力, 从而实现对合成的精确测量。这些方法可以用来测量酶的整体活性 (2.1 节) 或测量单个步骤的蔑氏参数 (注意以下步骤 2.1)。我们的实验室最近使用这些来表征以前进化的酶9以及合理设计的酶<sup class="xr...

dna 聚合进行准确和有效的 dna 合成, 是维持基因组完整性的关键。在不出错的情况下, 每秒合成数百个核苷酸的能力也使 DNA 聚合了分子生物学和生物技术的基本工具。然而, 这些属性也限制了 M DNA 基板的应用;一般而言, 天然 dna 聚合不能合成许多潜在的有价值的 M dna 基质, 可能是由于对使用非标准基质的高选择性压力在体内。许多小组开发了有针对性的进化方法来产生突变的 dna 聚合能够进行 M dna 合成1a,2,3,4,5;这些努力扩大了 DNA 的生物技术效用6,7,8。
为了评估突变 dna 聚合合成 M dna 的能力, 我们9,10, 其他11,12,13通常使用体外测量 dna聚合酶活性, 这是在这篇手稿中描述。在这些实验中, DNA 聚合是 co-incubated 与标记的底漆/模板复式和核苷三磷酸基板;用凝胶电泳法对产品进行评价。根据具体的实验问题, 突变的 DNA 聚合, 改良引物, 改良的模板, 或改性的核苷 triphosphates 可以使用, 使突变酶活性的系统生化评价。
从历史上看, 这些化验都依赖于5的放射性标签来跟踪 DNA 合成;最常见的是, 使用了32p 和33p。通常, 使用 T4 核苷酸激酶11实现标记。然而, 由于放射性标签的寿命有限, 成本相对较高, 安全处置, 我们组改用合成5的近红外荧光标记 DNA。使用相对低成本的近红外凝胶成像仪, 我们观察到了使用放射性标签 (未发表的结果) 之前研究的类似检测限制。我们已经成功地重现了过去的观测值9, 我们没有观察到与以前测量的速率常数 (未发布的结果) 有任何大的定量差异。
为了分析 dna 的大小, 因此, 我们依靠聚丙烯酰胺凝胶电泳方法, 在毛细管电泳15问世之前为桑格测序14 。分离或移动的距离可以用作分子量的测量;大格式, 垂直聚丙烯酰胺凝胶可以实现单核苷酸的分辨率, 使定量观察的 DNA 寡核苷酸的长度不等。
总的来说, 这些实验是一种用于聚合酶表征的健壮方法。由于反应的时间敏感性, 准备和护理是必要的, 以达到重现的结果。此外, 虽然丙烯酰胺凝胶是一种非常有效的方法来测量 dna 合成, 以及许多其他的 dna 修饰反应, 单核苷酸分辨率, 它可以在技术上具有挑战性。这里的协议有望使用户能够执行这些实验, 同时避免最常见的错误。

<table><tbody><tr><td>Tris HCl</td><td>Promega</td><td>H5123</td><td></td></tr><tr><td>Tris Base</td><td>Promega</td><td>H5131</td><td></td></tr><tr><td>MgCl&lt;sub&gt;2&lt;/sub&gt;</td><td>Fisher Scientific</td><td>BP214-500</td><td></td></tr><tr><td>Acetylated BSA</td><td>Promega</td><td>PR-R3961</td><td></td></tr><tr><td>KCl</td><td>Sigma</td><td>P4504</td><td></td></tr><tr><td>Dithiothreitol (i.e. DTT)</td><td>Research Products International</td><td>D11000</td><td></td></tr><tr><td>Ethylenediaminetetraacetic acid (i.e. EDTA) (0.5M solution)</td><td>Sigma</td><td>03690-100mL</td><td></td></tr><tr><td>Glycerol</td><td>Sigma</td><td>G5516</td><td></td></tr><tr><td>Formamide</td><td>Acros</td><td>AC42374-5000</td><td></td></tr><tr><td>Orange G</td><td>Sigma Aldrich</td><td>O3756</td><td></td></tr><tr><td>Bromophenol blue</td><td>Fisher Scientific</td><td>50-701-6973</td><td></td></tr><tr><td>dNTPs</td><td>Fisher Scientific</td><td>FERR0191</td><td></td></tr><tr><td>M-dNTPs (riboNTPs)</td><td>Fisher Scientific</td><td>45-001-341 (343, 345, 347)</td><td></td></tr><tr><td>M-dNTPs (all other modified NTPs)</td><td>TriLink Biotechnologies</td><td>assorted</td><td></td></tr><tr><td>primer 1</td><td>IDT DNA / TriLink Biotechnologies</td><td>Custom Syntheses</td><td>We use the IR700 dye which can be purchased as a custom synthesis. We typically purchase the oligonucleotides HPLC purified. Sequence is 5&amp;rsquo;-dTAATACGACTCACTATAGGGAGA</td></tr><tr><td>template 1</td><td>IDT DNA / TriLink Biotechnologies</td><td>Custom Syntheses</td><td>We typically purchase the oligonucleotides HPLC purified. Sequence is 5&amp;rsquo;-dCGCTAGGACGGCATTGGATCAGTCTCCCTATAGTGAGTCGTATTA</td></tr><tr><td>Acrylamide</td><td>Research Products International</td><td>A11405</td><td>38.67% acrylamide and 1.33% bis-acrylamide</td></tr><tr><td>Tris/Borate/EDTA (TBE) solid</td><td>Research Products International</td><td>T22020</td><td></td></tr><tr><td>Urea ultrapure</td><td>Research Products International</td><td>U20200</td><td></td></tr><tr><td>Gel tape</td><td>CBS Scientific</td><td>GT-72-10</td><td></td></tr><tr><td>Large white spring clamp polypropylene</td><td>CBS Scientific</td><td>GPC-0001</td><td></td></tr><tr><td>Ammonium persulfate (APS)</td><td>Fisher Scientific</td><td>BP179</td><td></td></tr><tr><td>Tetramethylethylenediamine (TEMED)</td><td>Fisher Scientific</td><td>BP15020</td><td></td></tr><tr><td>0.75 mm spacers</td><td>CBS Scientific</td><td>SGS-20-0740A</td><td></td></tr><tr><td>33x42 Notched Glass Plate Set</td><td>CBS Scientific</td><td>SGP33-040A</td><td></td></tr><tr><td>Wedge plate separator</td><td>CBS Scientific</td><td>WPS-100</td><td></td></tr><tr><td>Comb for gel electrophoresis</td><td>CBS Scientific</td><td>SG33-0734</td><td></td></tr><tr><td>Gel electrophoresis rig</td><td>CBS Scientific</td><td>SG-400-33</td><td></td></tr><tr><td>ultrapure water</td><td></td><td></td><td>we use a Milli-Q system from Millipore</td></tr><tr><td>DNA polymerases</td><td></td><td></td><td>we prepare these in our laboratory using published protocols.</td></tr></tbody></table>

dna polymerase activity assay using near-infrared fluorescent labeled dna visualized by acrylamide gel electrophoresis

对于任何酶, 都需要有健壮的、定量的方法来表征本地和工程酶。对于 dna 聚合, dna 合成可以用一种体外dna 合成方法进行表征, 然后采用聚丙烯酰胺凝胶电泳法。该方法的目的是量化合成的天然 dna 和修饰 dna (M-dna)。这些方法是特别有用的解决单核苷酸分解寡核苷酸, 使观察的个别步骤在酶寡核苷酸合成。这些方法已被用于评价的一系列生物化学和生物物理特性, 如测量的稳态速率常数的个别步骤的 dna 合成, 错误率的 dna 合成, 和 dna 结合的亲和力。通过修改后的核苷 triphosphates (NTP)、m-dna 和/或突变的 dna 聚合, 可以有效地评价基质 dna 聚合酶对的相对效用。在这里, 我们详细的分析本身, 包括必须作出的改变, 以适应非传统的底漆 dna 标记策略, 如近红外线荧光标记 dna。此外, 我们有详细的关键技术步骤, 丙烯酰胺凝胶浇注和运行, 这往往是技术上的挑战。

一个成功的聚丙烯酰胺凝胶分析的总体活动的定性描述 (在2.1 节,图 1) 和稳态动力学 (说明在说明中的 2.1,图 2) 显示。还显示了不成功的聚丙烯酰胺凝胶分析 (图 3)。
请注意, 没有商业可用的梯子或分子标记使用。有时, 我们将使用已知的聚合酶-基质结合物来创...

Watch this Scientific Journal Video about DNA Polymerase Activity Assay Using Near-infrared Fluorescent Labeled DNA Visualized by Acrylamide Gel Electrophoresis at JoVE.com

DNA Polymerase Activity Assay Using Near-infrared Fluorescent Labeled DNA Visualized by Acrylamide Gel Electrophoresis

用丙烯酰胺凝胶电泳法可视化近红外荧光标记 dna 聚合酶活性

本议定书描述了 dna 聚合酶合成的修饰 dna 通过观察的变化近红外荧光标记 dna 使用凝胶电泳和凝胶显像。丙烯酰胺凝胶用于高分辨率成像分离的短核酸, 这是根据大小不同的速率迁移。

For any enzyme, robust, quantitative methods are required for characterization of both native and engineered enzymes. For DNA polymerases, DNA synthesis ...

dna-polymerase-activity-assay-using-near-infrared-fluorescent-labeled

Research

JoVE Journal

Biochemistry

14.1K 次观看.  W.M. Keck Science Department of Claremont Mckenna, Pitzer, and Scripps College. 本议定书描述了 dna 聚合酶合成的修饰 dna 通过观察的变化近红外荧光标记 dna 使用凝胶电泳和凝胶显像。丙烯酰胺凝胶用于高分辨率成像分离的短核酸, 这是根据大小不同的速率迁移。

视频: DNA Polymerase Activity Assay Using Near-infrared Fluorescent Labeled DNA Visualized by Acrylamide Gel Electrophoresis

An Optimized Protocol for Electrophoretic Mobility Shift Assay Using Infrared Fluorescent Dye-labeled Oligonucleotides

Electrophoretic Mobility Shift Assays (EMSA) are an instrumental tool to characterize the interactions between proteins and their target DNA sequences. Radioactivity has been the predominant method of DNA labeling in EMSAs. However, recent advances in fluorescent dyes and scanning methods have prompted the use of fluorescent tagging of DNA as an alternative to radioactivity for the advantages of easy handling, saving time, reducing cost, and improving safety. We have recently used fluorescent EMSA (fEMSA) to successfully address an important biological question. Our fEMSA analysis provides mechanistic insight into the effect of a missense mutation, G73E, in the highly conserved HMG transcription factor SOX-2 on olfactory neuron type diversification. We found that mutant SOX-2G73E protein alters specific DNA binding activity, thereby causing olfactory neuron identity transformation. Here, we present an optimized and cost-effective step-by-step protocol for fEMSA using infrared fluorescent dye-labeled oligonucleotides containing the LIM-4/SOX-2 adjacent target sites and purified SOX-2 proteins (WT and mutant SOX-2G73E proteins) as a biological example.

We describe here an optimized protocol of fluorescent Electrophoretic Mobility Shift Assays (fEMSA) using purified SOX-2 proteins together with infrared fluorescent dye-labeled DNA probes as a case study to tackle an important biological question.

一种优化的协议电泳迁移分析使用红外荧光染料标记的寡核苷酸

Electrophoretic Mobility Shift Assays (EMSA) are an instrumental tool to characterize the interactions between proteins and their target DNA sequences. ...

科研

生物化学

DNA Staining Method Based on Formazan Precipitation Induced by Blue Light Exposure

DNA staining methods are very important for biomedical research. We designed a simple method that allows DNA visualization to the naked eye by the formation of a colored precipitate. It works by soaking the acrylamide or agarose DNA gel in a solution of 1x (equivalent to 2.0 &#181;M) SYBR Green I (SG I) and 0.20 mM nitro blue tetrazolium that produces a purple precipitate of formazan when exposed to sunlight or specifically blue light. Also, DNA recovery tests were performed using an ampicillin resistant plasmid in an agarose gel stained with our method. A larger number of colonies was obtained with our method than with traditional staining using SG I with ultraviolet illumination. The described method is fast, specific, and non-toxic for DNA detection, allowing visualization of biomolecules to the "naked eye" without a transilluminator, and is inexpensive and appropriate for field use. For these reasons, our new DNA staining method has potential benefits to both research and industry.

This method allows selective staining and quantification of DNA in gels by soaking the gel in a SYBR Green I/Nitro Blue Tetrazolium solution and then exposing it to sunlight or a blue light source. This produces a visible precipitate and requires almost no equipment, making it ideal for field use.

基于蓝光照射诱发甲沉淀的 DNA 染色方法

DNA staining methods are very important for biomedical research. We designed a simple method that allows DNA visualization to the naked eye by the ...

Nonradioactive Assay to Measure Polynucleotide Phosphorylation of Small Nucleotide Substrates

Polynucleotide kinases (PNKs) are enzymes that catalyze the phosphorylation of the 5&#39; hydroxyl end of DNA and RNA oligonucleotides. The activity of PNKs can be quantified using direct or indirect approaches. Presented here is a direct, in vitro approach to measure PNK activity that relies on a fluorescently-labeled oligonucleotide substrate and polyacrylamide gel electrophoresis. This approach provides resolution of the phosphorylated products while avoiding the use of radiolabeled substrates. The protocol details how to set up the phosphorylation reaction, prepare and run large polyacrylamide gels, and quantify the reaction products. The most technically challenging part of this assay is pouring and running the large polyacrylamide gels; thus, important details to overcome common difficulties are provided. This protocol was optimized for Grc3, a PNK that assembles into an obligate pre-ribosomal RNA processing complex with its binding partner, the Las1 nuclease. However, this protocol can be adapted to measure the activity of other PNK enzymes. Moreover, this assay can also be modified to determine the effects of different components of the reaction, such as the nucleoside triphosphate, metal ions, and oligonucleotides.

This protocol describes a nonradioactive assay to measure kinase activity of polynucleotide kinases (PNKs) on small DNA and RNA substrates.

测量小核苷酸基质的多核苷酸磷酸化的非放射性测定

Polynucleotide kinases (PNKs) are enzymes that catalyze the phosphorylation of the 5&#39; hydroxyl end of DNA and RNA oligonucleotides. The activity of ...

使用纳米荧光素酶报告基因检测定量 DNA 双链断裂修复

Assessment of DNA Double Strand Break Repair Activity Using High-throughput and Quantitative Luminescence-Based Reporter Assays

The repair of DNA double strand breaks (DSBs) is crucial for the maintenance of genome stability and cell viability. DSB repair (DSBR) in cells is mediated through several mechanisms: homologous recombination (HR), non-homologous end joining (NHEJ), microhomology-mediated end joining (MMEJ), and single strand annealing (SSA). Cellular assays are essential to measure the proficiency and modulation of these pathways in response to various stimuli.
Here, we present a suite of extrachromosomal reporter assays that each measure the reconstitution of a nanoluciferase reporter gene by one of the four major DSBR pathways in cells. Upon transient transfection into cells of interest, repair of pathway-specific reporter substrates can be measured in under 24 h by the detection of Nanoluciferase (NanoLuc) luminescence.
These robust assays are quantitative, sensitive, titratable, and amenable to a high-throughput screening format. These properties provide broad applications in DNA repair research and drug discovery, complementing the currently available toolkit of cellular DSBR assays.

作者聚焦：开发针对 DNA 损伤反应的新型抗癌疗法

We present protocols for the assessment of double strand break repair pathway proficiency in cells using a suite of luminescence-based extrachromosomal reporter substrates.

使用基于高通量和定量发光的报告基因分析评估 DNA 双链断裂修复活性

The repair of DNA double strand breaks (DSBs) is crucial for the maintenance of genome stability and cell viability. DSB repair (DSBR) in cells is ...

癌症研究

<meta charset="utf8"></head><body>用于鉴别性 RNA 和 DNA 分析的精密 DNAzyme 纳米机器

DNAzyme 10-23 - Based Nanomachines for Nucleic Acid Recognition

DNAzyme-based nanomachines (DNM) for the detection of DNA and RNA sequences (analytes) are multifunctional structures made of oligonucleotides. Their functions include tight analyte binding, highly selective analyte recognition, fluorescent signal amplification by multiple catalytic cleavages of a fluorogenic reporter substrate, and fluorogenic substrate attraction for an increase in sensor response. Functional units are attached to a common DNA scaffold for their cooperative action. The RNA-cleaving 10-23 DNMs feature&#160;improved sensitivity in comparison with non-catalytic hybridization probes. The stability of the DNM and the increased chances of substrate recognition are provided&#160;by a double-stranded DNA fragment,&#160;a tile. DNM can differentiate two analytes with a single nucleotide difference in a folded RNA and a double-stranded DNA and detect analytes at concentrations ~1000 times lower than other protein-free hybridization probes. This article presents the concept behind the diagnostic potential of DNA-nanomachine activity and overviews DNM design, assembly, and application&#160;in nucleic acid detection assays.

<meta charset="utf8"></head><body>作者聚焦:DNA 纳米传感器的进步 – 解决分子检测中的灵敏度和选择性挑战

The DNAzyme-based nanomachines can be used for highly selective and sensitive detection of nucleic acids. This article describes a detailed protocol for the design of DNAzyme-based nanomachines with a 10-23 core using free software and their application in the detection of an Epstein-Barr virus fragment as an example.

DNAzyme 10-23 - 用于核酸识别的基于纳米机器

DNAzyme-based nanomachines (DNM) for the detection of DNA and RNA sequences (analytes) are multifunctional structures made of oligonucleotides. Their ...

使用合成寡核苷酸的 DNA 酶检测

Using Modified Synthetic Oligonucleotides to Assay Nucleic Acid-Metabolizing Enzymes

The availability of a range of modified synthetic oligonucleotides from commercial vendors has allowed the development of sophisticated assays to characterize diverse properties of nucleic acid metabolizing enzymes that can be run in any standard molecular biology lab. The use of fluorescent labels has made these methods accessible to researchers with standard PAGE electrophoresis equipment and a fluorescent-enabled imager, without using radioactive materials or requiring a lab designed for the storage and preparation of radioactive materials, i.e., a Hot Lab. The optional addition of standard modifications such as phosphorylation can simplify assay setup, while the specific incorporation of modified nucleotides that mimic DNA damages or intermediates can be used to probe specific aspects of enzyme behavior. Here, the design and execution of assays to interrogate several aspects of DNA processing by enzymes using commercially available synthetic oligonucleotides are demonstrated. These include the ability of ligases to join or nucleases to degrade different DNA and RNA hybrid structures, differential cofactor usage by the DNA ligase, and evaluation of the DNA-binding capacity of enzymes. Factors to consider when designing synthetic nucleotide substrates are discussed, and a basic set of oligonucleotides that can be used for a range of nucleic acid ligase, polymerase, and nuclease enzyme assays are provided.

作者聚焦：表征来自极端微生物和常见病原体的新型酶以了解 DNA 修复和复制

Here, a protocol for assaying nucleic acid metabolizing enzymes is presented, using examples of ligase, nuclease, and polymerase enzymes. The assay utilizes fluorescently labeled and unlabeled oligonucleotides that can be combined to form duplexes mimicking RNA and/or DNA damages or pathway intermediates, allowing for the characterization of enzyme behavior.

用丙烯酰胺凝胶电泳法可视化近红外荧光标记 dna 聚合酶活性

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一种优化的协议电泳迁移分析使用红外荧光染料标记的寡核苷酸

基于蓝光照射诱发甲沉淀的 DNA 染色方法

测量小核苷酸基质的多核苷酸磷酸化的非放射性测定

使用基于高通量和定量发光的报告基因分析评估 DNA 双链断裂修复活性

使用基于高通量和定量发光的报告基因分析评估 DNA 双链断裂修复活性

使用基于高通量和定量发光的报告基因分析评估 DNA 双链断裂修复活性

DNAzyme 10-23 - 用于核酸识别的基于纳米机器

DNAzyme 10-23 - 用于核酸识别的基于纳米机器

DNAzyme 10-23 - 用于核酸识别的基于纳米机器

使用修饰的合成寡核苷酸检测核酸代谢酶