PrimRglo: A multiplexable quantitative real-time polymerase chain reaction system for nucleic acid detection

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• 作者：Richard Lai^a ; ^b ; Fang Liang^b ; Darnley Pearson^a ; Graeme Barnett^a ; David Whiley^c ; Theo Sloots^c ; Ross T. Barnard^b ; ^{rossbarnard@uq.edu.au} ; Simon R. Corrie^d
• 关键词：Quantitative ; Real time ; Polymerase chain reaction ; Multiplex
• 刊名：Analytical Biochemistry
• 出版年：2012
• 期刊代码：93_00032697
• 类别：imm
• 出版时间：15 March, 2012
• 卷：422
• 期：2
• 页码：89-95
• 文件大小：462 K

摘要

We report the development of a new real-time polymerase chain reaction (PCR) detection system that uses oligonucleotide 鈥渢agged鈥?PCR primers, a fluorophore-labeled 鈥渦niversal鈥?detection oligonucleotides, and a complementary quenching oligonucleotide. The fluorescence signal decreases as PCR product accumulates due to the increase in detection/quencher hybrid formation as the tagged primer is consumed. We use plasmids containing the influenza A matrix gene and the porA and ctrA genes of Neisseria meningitidis as targets for developing the system. Cycle threshold (C_t) values were generated, and the sensitivity of the new system (dubbed 鈥淧rimRglo鈥? compared favorably with the commonly used SYBR green and Taqman detection systems and, unlike the latter system, does not require the design of a new dual-labeled detection oligonucleotide for each new target sequence.