Frech Christian, Breuer Karin, Ronacher Bernhard, Kern Thomas, Sohn Christof, Gebauer Gerhard
University of Applied Sciences, Softwarepark 11, 4232 Hagenberg, Austria.
Comput Methods Programs Biomed. 2009 May;94(2):152-60. doi: 10.1016/j.cmpb.2008.12.007. Epub 2009 Feb 6.
Due to recent advances in genome sequencing, the detection of pathogens by DNA signatures, i.e. by oligonucleotide sequences that uniquely identify a specific genome, is becoming increasingly popular in modern clinical diagnostics. However, currently available screening methods, such as PCR and microarrays, lack multiplexing and sensitivity, respectively. Solid-phase amplification (SPA) is an emerging approach with the potential to overcome these limitations. SPA-based diagnostic assays require both pathogen-specific and compatible primer pairs for many, often closely related pathogens. Currently, none of the available tools supports an automated design of such primer sets, making it an iterative, labor-intensive, and often difficult procedure. Here we describe hybseek, a Web interface for efficient design of both pathogen-specific and compatible primer pairs for DNA-based diagnostic multi-analyte assays. hybseek achieves pathogen-specificity by selecting only candidates with unique 3(') subsequence, and the degree of this uniqueness is quantitatively expressed by a specificity score. qPCR experimental data confirm the feasibility of our design strategy. The service is freely available at https://www.hybseek.com.
由于基因组测序的最新进展，通过DNA特征（即通过唯一识别特定基因组的寡核苷酸序列）检测病原体在现代临床诊断中越来越受欢迎。然而，目前可用的筛查方法，如聚合酶链反应（PCR）和微阵列，分别缺乏多重检测能力和灵敏度。固相扩增（SPA）是一种新兴方法，有潜力克服这些局限性。基于SPA的诊断检测需要针对许多通常密切相关的病原体设计病原体特异性且兼容的引物对。目前，现有的工具都不支持自动设计此类引物组，这使得该过程成为一个反复进行、劳动强度大且往往困难的程序。在此，我们描述了hybseek，这是一个用于高效设计基于DNA的诊断多分析物检测中病原体特异性且兼容引物对的网络界面。hybseek通过仅选择具有独特3′子序列的候选引物来实现病原体特异性，这种独特性程度由特异性评分定量表示。定量聚合酶链反应（qPCR）实验数据证实了我们设计策略的可行性。该服务可在https://www.hybseek.com免费获取。
