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MicroRNA-derived fragment length polymorphism assay.

ABSTRACT: MicroRNA (miRNA) studies are experiencing a transition from basic research applications to clinical applications. However, the lack of reliable and sensitive miRNA detection methods has become a bottleneck in the process. Here, we report an absolute quantification method based on the competitive PCR amplification of specific miRNAs and synthetic RNA spike-ins in a single reaction. RNA spike-ins are quantified as dynamic RNA copy number standards and are used to measure selected miRNAs free from the effects of intra-assay variables, including those from individual sample sources. Combined with the size differentiation power of capillary electrophoresis, the content of miRNAs was reproducibly measured, with verifiable detection limits of 10-46 copies over 5-log detection ranges. The direct measurements of miRNAs from 168 human serum samples and their considerable value as a diagnostic for bronchopneumonia and bronchiolitis demonstrate the potential of the assay in clinical applications.

SUBMITTER: Xie X

PROVIDER: S-EPMC4366852 | biostudies-other | 2015

REPOSITORIES: biostudies-other

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MicroRNA-derived fragment length polymorphism assay.

Xie Xiaoping X Tang Fang F Yang Zhao Z Zhang Yaoyi Y Feng Zihao Z Yang Yu Y Wu Xiujin X Zhang Feifei F Zhu Jie J Xu Kai K

Scientific reports 20150320

MicroRNA (miRNA) studies are experiencing a transition from basic research applications to clinical applications. However, the lack of reliable and sensitive miRNA detection methods has become a bottleneck in the process. Here, we report an absolute quantification method based on the competitive PCR amplification of specific miRNAs and synthetic RNA spike-ins in a single reaction. RNA spike-ins are quantified as dynamic RNA copy number standards and are used to measure selected miRNAs free from ...[more]

PMID: 25790971

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