PCR技术

Extrachromosomal Elements-Plasmids (Michael Blaber)What's plasmid? Find answer here. It provides background information on the features of plasmid selection marker and more... Rescuing Integrated ...

Recommended Reaction Conditions: Initial Conditions: Initial denaturation at start: 92 - 97oC for 3 - 5 min. If you denature at 97oC denature sample only; add rest of mix after reaction colls to annea ...

Labelling PCR Products with Digoxigenin PCR products may be very conveniently labelled with digoxigenin-11-dUTP (Boehringer-Mannheim) by incorporating the reagent to 10-35% final effective dTTP concen ...

"Hot Start" PCR: In certain circumstances one wishes to avoid mixing primers and target DNA at low temperatures in the presence of Taq polymerase: Taq pol is almost as efficient as Klenow pol at 37oC; ...

Cloning PCR Products T-A Cloning Strategy: Taq and other polymerases seem to have a terminal transferase activity which results in the non-templated addition of a single nucleotide to the 3'-ends of P ...

・ Plasmid Midi-preps (NWSFC)Diatomaceous Earth-based midi-prep. This procedure is the method of choice for isolating double stranded plasmid-based templates for the Sequenase Dye-Labeled Termi ...

・ Standard (alkaline lysis) Mini-Prep (Goldberg Lab)Standard protocol for mini-prep and recipe for solution I II and III.Alkaline Lysis Minipreps of Plasmid or Cosmid DNA (Donis Keller Lab)Pla ...

・ Plasmid Mini and Maxi Prep Methods (Gimila Lab) ・ Maxi-preps and all media solutions (NWFSC)Isolation of cosmid plasmid and P1 DNAs via diatomaceous earth-based Maxi Midi and Mini-p ...

Preparation of Cosmid DNAPreparation of Cosmid DNA from 50 ml Cultures (Donis Keller Lab)Cosmid vectors containing foreign DNA inserts are known to replicate more slowly than small plasmids in culture ...

把PCR产物克隆到载体上常见有3种做法：1.直接克隆到T载体（或者U载体上）2.引物设计时引入酶切位点，PCR产物酶切后直接克隆到目标载体上3.将平末端PCR产物直接克隆到平末端酶切载体上。 方法3主要是针对高保真的聚合酶比如Stratagene公司的Pfu酶，New England Biolabs公司的Vent酶，以及QIAGEN公司新出的同时具有热启动功能的高保真ProofStart酶 ...

PCR条件的优化是一个麻烦耗时的过程，涉及到温度、时间、镁离子、引物、dNTP、Taq酶、模板等多个因素的调整。一般来说利用热启动，比如Platinum Taq DNA聚合酶（Invitrogen）可以达到更高的特异性，降低对镁离子浓度的依赖，并且有利于提高“问题模板”的产量。然而，传统的PCR条件优化策略经常对含有稳定二级结构的DNA序列无效，因为这些序列难以变性，并且在退火过程中影响引物与模板 ...

肖生祥 （西安医科大学第二临床医学院皮肤科，710004）　　 开展 PCR 应具有一定的条件。需要一个正规的 pCR 实验室，采集标本、核酸提取、扩增及产物分析应在不同的房间进行；需有严密的防污染措施、假阳性和假阴性结果的质控等。实验者应具有一定的分子生物学理论知识及实验技能，能够及时发现和纠正实验中的问题。目前，我国在 PCR 应用中尤其是在临床诊断中存在问题较多，主要表现为： PCR 试剂的 ...

Simple ja;ascript Oligo CalculatorOligo CalculatorEnter Oligo Sequence in Box Length Melting Temperature (Tm) °C %GC content Molecular Weight: daltons (g/M) OD of 1 is equal to picoMolar. ...

PCR产物的克隆 PCR产物克隆大致分为两类，即平头连接和粘头连接。 平头连接是将制备好的平头载体和补平或削平的PCR产物直接进行连接。载体可用EcoR V或Sma I切成平头；PCR产物纯化后，可以在22℃用DNA聚合酶I作用30min（利用该酶所具有的3’→5’外切酶活性和5’→3’的聚合酶活性）。如果要求不高，PCR产物也可不加处理。如果使用Stratagene公司的pfu DNA聚合 ...

　 聚合酶链反应-单链构象多态性分析（Single Strand Conformation Polymorphism Analysis of Polymerase Chain Reaction Products PCR-SSCP）是近年来发展起来的一种基因分析方法。 　PCR-SSCP分析的基本程序为：首先PCR扩增特定靶序列，然后将扩增产物变性为单链，进行非变性聚丙烯酰胺凝胶电泳。在不含 ...

逆转录-聚合酶链反应 (Reverse Transcription-Polymerase Chain Reaction，RT-PCR)的原理是：提取组织或细胞中的总RNA，以其中的mRNA作为模板，采用Oligo（dT）或随机引物利用逆转录酶反转录成cDNA。再以cDNA为模板进行PCR扩增，而获得目的基因或检测基因表达。RT-PCR使RNA检测的灵敏性提高了几个数量级，使一些极为微量RNA样品分 ...

相关文件如下： 点击浏览该文件PCR方法相关产品:电泳设备紫外设备普通PCR仪定量PCR仪PCR/RT-PCR/qPCR试剂PCR引物 PCR试剂PCR对照特异性PCR试剂盒PCR克隆试剂盒RNA RNase检测/去除 RT-PCR试剂 RT-PCR标准品 定量PCR试剂 定量PCR标记 总RNA分离纯化盒PCR产物纯化核酸酶 聚合酶 反转录酶 ...

Various authors recommend DMSO and glycerol to improve amplification efficiency (higher amount of product) and specificity (no unspecific products) of PCR when used in concentrations varying between 5 ...

A variety of PCR additives and enhancing agents have been used to increase the yield specificity and consistency of PCR reactions. Whilst these additives may have beneficial effects on some amplificat ...

Beforehand.choose empirically the annealing temperature of primers; make sure you have enough PCR-plates Q-covers and sealing tape; prepare the stocks: PCR-buffer 5M Betaine - with some excess dNTPs p ...