MicroRNAs, serve as critical gene expression regulators at the transcriptional and post-transcriptional levels, have also been found as important blood-based biomarkers for early detection of cancers. However, their current in vitro detection methods are relatively complex, costly and low sensitive. Our project attempts to establish a novel in vitro microRNA detection system which is rapid, efficient, sensitive and specific. In this system, CRISPR-Cas9 technique is modified to integrate with split-HRP reporting systems. The advanced rolling circle amplification technology and cell-free expression system are also involved and optimized. This system may ideally be compatible for the detection of various series of small non-coding RNAs. To our knowledge, we are the first to use the CRISPR-Cas9 system as a small non-coding RNA monitor in vitro. Its establishment and further development might provide a new approach for rapid and low-cost cancer screening, virus detection and curative efficacy assessment.