期刊
BIOSENSORS & BIOELECTRONICS
卷 74, 期 -, 页码 778-785出版社
ELSEVIER ADVANCED TECHNOLOGY
DOI: 10.1016/j.bios.2015.06.054
关键词
Paper-based nucleic acid diagnostics; Universal primer mediated asymmetric PCR; Multiplex detection; Genotyping; Point-of-care application
类别
资金
- National Basic Research Program of China [2010CB732602]
- National Natural Science Foundation of China, China [21475048]
- National Science Fund for Distinguished Young Scholars of Guangdong Province [2014A030306008]
- Program of the Pearl River Young Talents of Science and Technology in Guangzhou, China [2013J2200021]
Traditionary multiplex asymmetric polymerase chain reaction (PCR) can be applied to detect multiplex target organisms simultaneously, but complex optimizations of primer concentrations and staggered additions of primers are required to achieve equal amplification of multiplex genes. To overcome this shortcoming, we propose a novel method based on multiplex asymmetric PCR and paper-based nucleic acid diagnostics (PBNAD). In the asymmetric PCR, a universal primer was introduced to break the bottlenecks of low sensitivity and self-inhibition among different sets of primers. Amplification using the novel multiplex asymmetric PCR boosted the quantity of single-stranded amplicons, and the amplified products contained the same sequence at the 5' end. Therefore, only one gold nanoparticle-based signal probe was needed for the simultaneous detection of three genes using the PBNAD platform, and the detection signals could be observed with the naked eye. With this highly efficient, novel multiplex asymmetric PCR, as little as 1 pg/mu L genomic DNA can be detected. This method can also be applied to genotyping for reliable epidemiological investigations. This proof-of-concept study highlights the potential of the PBNAD platform for cost- and labor-effective applications in the detection of pathogenic bacteria. (C) 2015 Elsevier B.V. All rights reserved.
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