Kumar Anil, Chordia Nikita
School of Biotechnology, Devi Ahilya University, Khandwa Rd., Indore, 452001, India,
Methods Mol Biol. 2015;1275:143-51. doi: 10.1007/978-1-4939-2365-6_10.
Polymerase chain reaction (PCR) is an enzymatic reaction whose efficiency and sensitivity largely depend on the efficiency of the primers that are used for the amplification of a concerned gene/DNA fragment. Selective amplification of nucleic acid molecules initially present in minute quantities provides a powerful tool for analyzing nucleic acids. In silico method helps in designing primers. There are various programs available for PCR primer design. Here we described designing of primers using web-based tools like "Primer3" and "Web Primer". For designing the primer, DNA template sequence is required that can be taken from any of the available sequence databases, e.g., RefSeq database. The in silico validation can be carried out using BLAST tool and Gene Runner software, which check their efficiency and specificity. Thereafter, the primers designed in silico can be validated in the wet lab. After that, these validated primers can be synthesized for use in the amplification of concerned gene/DNA fragment.
聚合酶链反应（PCR）是一种酶促反应，其效率和灵敏度在很大程度上取决于用于扩增相关基因/DNA片段的引物的效率。对最初以微量存在的核酸分子进行选择性扩增为分析核酸提供了一个强大的工具。计算机辅助方法有助于引物设计。有各种可用于PCR引物设计的程序。在这里，我们描述了使用诸如“Primer3”和“Web Primer”等基于网络的工具进行引物设计。为了设计引物，需要DNA模板序列，其可以取自任何可用的序列数据库，例如RefSeq数据库。可以使用BLAST工具和Gene Runner软件进行计算机辅助验证，以检查它们的效率和特异性。此后，可以在湿实验室中验证在计算机上设计的引物。之后，可以合成这些经过验证的引物，用于扩增相关基因/DNA片段。
