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所在平台: Udemy |
课程主页: https://www.udemy.com/course/learn-dna-primer-design-for-polymerase-chain-reaction/
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课程名称：学习聚合酶链反应的DNA引物设计 课程概述：本生物信息学课程将教您如何为聚合酶链反应（PCR）设计DNA引物。Primer BLAST工具在提供目标模板和两个引物后，仅执行特异性检查。您可以使用我们的引物设计工具，设计用于单个或多个插入克隆的引物，或者进行定点突变实验（插入、缺失、替代）。Primer3是一个计算机程序，可以为多种应用建议PCR引物，例如创建用于辐射杂交图谱的序列标签位点（STS），或放大序列以发现单核苷酸多态性（SNP）。 该课程将详细介绍PCR的步骤，包括变性、退火和延伸，以及引物设计的规范。好的引物设计应确保GC含量在40%到60%之间，并且引物的3'末端建议以G或C结束以促进结合。PCR引物的理想长度通常在18到30个碱基之间。引物的熔解温度（Tm）应在65°C到75°C之间，并且相互之间应保持在5°C之内。引物的特异性通常取决于长度和退火温度。引物越短，与目标结合或退火的效率越高。但引物不应过长（30个碱基以上）或过短。过短的引物会产生不准确和非特异性的DNA扩增产物，而过长的引物则会导致杂交速率减慢。此外，还需避免引物间的连接，这会产生引物二聚体，干扰扩增过程。 该课程为学习PCR引物设计提供了全面的知识和实用技能，对生物研究和实验室工作具有重要意义。
In this Bioinformatics course you will be find out how to DNA Primer Design for polymerase chain reaction. Primer BLAST performs only a specificity check when a target template and both primers are provided. Design primers for single- or multi-insert cloning or for your site-directed mutagenesis experiment (insertion, deletion, replacement) with our primer design tool. Primer3 is a computer program that suggests PCR primers for a variety of applications, for example to create STSs (sequence tagged sites) for radiation hybrid mapping, or to amplify sequences for single nucleotide polymor- phism discoveryPolymerase chain reaction (PCR) stepsDenaturingAnnealingExtensionSpecification of Primer DesignAim for the GC content to be between 40 and 60% with the 3' of a primer ending in G or C to promote bindingA good length for PCR primers is generally around 18-30 bases.Try to make the melting temperature (Tm) of the primers between 65°C and 75°C, and within 5°C of each otherA good length for PCR primers is generally around 18-30 bases. Specificity usually is dependent on length and annealing temperature. The shorter the primers are, the more efficiently they will bind or anneal to the target.However, a primer should not be too long 30-mer primers) or too short. Short primers produce inaccurate, nonspecific DNA amplification product, and long primers result in a slower hybridizing rate..One also needs to avoid primer-primer annealing which creates primer dimers and disrupts the amplification process
