Bielawski J P, Noack K, Pumo D E
Hofstra University, Hempstead, NY, USA.
Biotechniques. 1995 May;18(5):856-60.
Randomly amplified polymorphic DNA (RAPD) is a modification of PCR that uses short, randomly generated primers to amplify genomic DNA. Generally, many bands of mixed intensity (i.e., strong, faint, fuzzy or sharp) are generated with each primer. Mixed-intensity bands are inherent with the RAPD technique because (i) the target DNAs are undefined, (ii) one or more copies of the target DNA may exist per genome and (iii) the percentage of hybridization of primer to target may vary. The problem of mixed-intensity band exacerbates the well-known sensitivity of PCRs to reagent and template concentrations, pH and other reaction parameters. These complications have discouraged many investigators from using RAPD. Our goal was to optimize the RAPD amplification conditions for vertebrate DNA. We present the optimized protocol along with an experimental strategy for obtaining reproducible, interpretable RAPD banding patterns in vertebrates.
随机扩增多态性DNA（RAPD）是聚合酶链式反应（PCR）的一种改良方法，它使用短的、随机生成的引物来扩增基因组DNA。通常，每个引物会产生许多强度各异的条带（即强、弱、模糊或清晰）。强度各异的条带是RAPD技术所固有的，原因如下：（i）靶DNA不明确；（ii）每个基因组中可能存在一个或多个靶DNA拷贝；（iii）引物与靶标的杂交百分比可能不同。强度各异的条带问题加剧了PCR对试剂和模板浓度、pH值及其他反应参数的众所周知的敏感性。这些复杂情况使得许多研究人员不愿使用RAPD。我们的目标是优化脊椎动物DNA的RAPD扩增条件。我们展示了优化后的方案以及一种实验策略，用于在脊椎动物中获得可重复、可解释的RAPD条带模式。
