Highly-sensitive multicomponent simultaneous detection method and system of miRNAs in single cells on basis of droplet microfluidic technology

Abstract

The invention provides a highly-sensitive multicomponent simultaneous detection method and system of microRNAs in single cells on the basis of a droplet microfluidic technology, and belongs to the technical field of biological detection and molecular biology. Compared with the prior art, the efficient cell processing capacity is achieved, 100 cells can be captured per second, and 13,000 cells canbe detected per hour. By means of secondary isothermal amplification in droplets, the amplification reaction rate and detection sensitivity are improved, multiple down-regulated expression microRNAs in a single cell can be analyzed. Isolation space in droplets can effectively prevent degradation of the microRNAs and ensure the accuracy of detection, and the method and system have good practical application value.

Description

technical field [0001] The invention belongs to the technical field of biological detection and molecular biology, and in particular relates to a highly sensitive multi-component simultaneous detection method and system for miRNA in a single cell based on droplet microfluidic technology. Background technique [0002] The information disclosed in this background section is only intended to increase the understanding of the general background of the present invention, and is not necessarily taken as an acknowledgment or any form of suggestion that the information constitutes the prior art already known to those skilled in the art. [0003] MicroRNA (miRNA) is a kind of endogenous short-chain RNA with a length of about 20-24 nucleotides, which plays a variety of important biological functions through the regulation of protein-coding gene mRNA, such as cell growth and differentiation , apoptosis, and proliferation. Abnormal expression of miRNA in cells is associated with many d...

AI Technical Summary

Problems solved by technology

With the gradual emergence of the value of miRNA in biology and clinical medicine, highly selective and sensitive miRNA analysis methods in cells have been continuously developed. At present, most of the detection methods for miRNA are based on the analysis of cell homogenate, which can neither Adapt to the processing and detection requirements of micro-cell samples, and it is difficult to avoid the potential degradation of miRNA by enzymes caused by exposure to the environment after cell membrane rupture

Although the sensitivity of some miRNA detection methods can reach single-cell or even single-molecule level, there are few studies on real miRNA single-cell quantitative analysis. It has been reported that a single cell lysate can be dispersed to 20,000 droplets and then amplified. , realized the quantitative detection of miRNA single cells, but the inventors found that the difficulty of single cell manipulation led to low throughput of sample processing and single cell analysis, which restricted the study of cell heterogeneity of miRNAs expression, and further limited its use in Practical Applications in Biomedicine

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