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首页> 外文期刊>Biochemistry and molecular biology education >Undergraduate Virology Exercises Demonstrate Conventional and Real-time PCR Using Commercially Available HIV Primers and Noninfectious Target
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Undergraduate Virology Exercises Demonstrate Conventional and Real-time PCR Using Commercially Available HIV Primers and Noninfectious Target

机译：大学生病毒学练习展示了使用市售HIV引物和非感染性靶标进行的常规PCR和实时PCR

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摘要

It is an extraordinary challenge to offer an undergraduate laboratory course in virology that teaches hands-on, relevant molecular biology techniques using nonpathogenic models of human virus detection. To our knowledge, there exists no inexpensive kits or reagent sets that are appropriate for demonstrating real-time PCR (RT-PCR) in an undergraduate laboratory course in virology. Here we describe simple procedures for student exercises that demonstrate the PCR detection of an HIV target nucleic acid. Our procedures combine a commercially available kit for conventional PCR with a modification for RT-PCR using the same reagents in the kit, making it possible for an instructor with access to a LightCycler~R instrument to implement a relevant student exercise on RT-PCR detection of HIV nucleic acid targets. This combination of techniques is useful for demonstrating and comparing conventional PCR amplification and detection with agarose gel electrophoresis, with real-time PCR over a series of three laboratory periods. The series of laboratory periods also is used to provide the foundation for teaching the concept of PCR primer design, optimization of PCR detection systems, and introduction to nucleic acid queries using NCBI-BLAST to find and identify primers, amplicons, and other potential amplification targets within the HIV viral genome. The techniques were successfully implemented at the Biology 364 undergraduate virology course at the University of Scranton during the Fall 2008 semester. The techniques are particularly targeted to students who intend to pursue either postgraduate technical employment or graduate studies in the molecular life sciences.
机译：提供一门病毒学本科实验室课程是一个非凡的挑战，该课程使用人类病毒检测的非致病性模型教授动手的相关分子生物学技术。据我们所知，没有便宜的试剂盒或试剂盒适合在病毒学本科实验室课程中展示实时PCR（RT-PCR）。在这里，我们描述了用于学生练习的简单程序，这些程序演示了HIV靶核酸的PCR检测。我们的程序结合了市售的常规PCR试剂盒和使用试剂盒中相同试剂进行RT-PCR修饰的试剂盒，使教师可以使用LightCycler〜R仪器实施有关RT-PCR检测的学生练习HIV核酸靶标。这种技术组合可用于演示和比较常规PCR扩增和琼脂糖凝胶电泳检测，以及在三个实验室周期中进行的实时PCR。该系列实验室时间还用于为教授PCR引物设计，优化PCR检测系统以及使用NCBI-BLAST查找和鉴定引物，扩增子和其他潜在扩增靶点的核酸查询简介提供基础在HIV病毒基因组中。该技术已在2008年秋季学期的斯克兰顿大学的生物学364本科病毒学课程中成功实施。该技术特别针对打算从事分子生命科学的研究生技术工作或研究生课程的学生。

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