A method for generating sticky-end PCR products which facilitates unidirectional cloning and the one-step assembly of complex DNA constructs.

Walker A; Taylor J; Rowe D

首页> 外文期刊>Plasmid: An International Journal Devoted to Extrachromosomal Gene Systems >A method for generating sticky-end PCR products which facilitates unidirectional cloning and the one-step assembly of complex DNA constructs.

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A method for generating sticky-end PCR products which facilitates unidirectional cloning and the one-step assembly of complex DNA constructs.

机译：一种产生粘性末端PCR产物的方法，其促进单向克隆和复杂DNA构建体的一步组装。

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We have developed and tested a method for the restriction enzyme-independent generation of sticky-end PCR products. The method is suitable for use with a proof-reading polymerase such as pfu, or any other heat-stable polymerase which produces a blunt-end product. The technique can be used to achieve unidirectional cloning of PCR products with an efficiency greater than 90%. Because the sequences of the sticky ends are defined by the user and potentially can be of any length, the method can also be exploited for the one-step construction of recombinant plasmids from multiple functional cassettes, without the use of restriction enzymes.

机译：我们已经开发并测试了一种不依赖于限制性内切酶的粘性末端PCR产物的生成方法。该方法适合与校对聚合酶（例如pfu）或产生平末端产物的任何其他热稳定聚合酶一起使用。该技术可用于以90％以上的效率实现PCR产物的单向克隆。由于粘性末端的序列是由用户定义的，并且可能具有任意长度，因此该方法还可用于从多个功能盒一步构建重组质粒，而无需使用限制酶。

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《Plasmid: An International Journal Devoted to Extrachromosomal Gene Systems》 |2008年第3期|共8页
作者
Walker A; Taylor J; Rowe D;
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正文语种 eng
中图分类医学遗传学;
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1. A method for generating sticky-end PCR products which facilitates unidirectional cloning and the one-step assembly of complex DNA constructs. [J] . Walker A, Taylor J, Rowe D Plasmid: An International Journal Devoted to Extrachromosomal Gene Systems . 2008,第3期

机译：一种产生粘性末端PCR产物的方法，其促进单向克隆和复杂DNA构建体的一步组装。
2. Turbo cloning: a fast, efficient method for cloning PCR products and other blunt-ended DNA fragments into plasmids [J] . A.Christopher Boyd Nucleic acids research . 1993,第4期

机译：Turbo克隆：一种快速，有效的方法，可将PCR产物和其他平末端DNA片段克隆到质粒中
3. Cloning and Assembly of PCR Products Using Modified Primers and DNA Repair Enzymes [J] . BioTechniques . 1997,第5期

机译：使用修饰的引物和DNA修复酶克隆和组装PCR产品
4. Quantitative Nucleic Acid Analysis Using Digital PCR: Absolute Counting of DNA (Examples Including Liquid Biopsies in Cancer) and RNA (mRNA and miRNA Using One-Step RT-dPCR) [C] . Michael Samuels, Leonora Balaj, Ben Ho Park, Molecular Medicine TRI-CON. . 2014

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5. Use of PCR Cloning Combined with DNA Barcoding to Identify Fish in a Mixed-Species Product [D] . Silva, Anthony J. 2019

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6. PCR Duplication: A One-Step Cloning-Free Method to Generate Duplicated Chromosomal Loci and Interference-Free Expression Reporters in Yeast [O] . Florian Huber, Matthias Meurer, Daria Bunina, -1

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7. HomeRun Vector Assembly System: a flexible and standardized cloning system for assembly of multi-modular DNA constructs. [O] . Ming V Li, Dip Shukla, Brian H Rhodes, 2014

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8. Methods, Systems, and Computer Program Products for Identifying Computer Program Source Code Constructs. [R] . Smith, J. M., Stotts, P. D. 2004

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A method for generating sticky-end PCR products which facilitates unidirectional cloning and the one-step assembly of complex DNA constructs.

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