Nanoprobe for real-time parallel detection of content of various mRNA in living cells

Abstract

The invention belongs to the technical field of nucleic acid detection and relates to a nanoprobe for real-time parallel detection of content of various mRNA in living cells. The nanoprobe comprises a penetrating peptide, a quantum dot, capture probes, reporter probes and gold nanoparticles. The quantum dot serving as a donor is connected with the capture probes; the gold nanoparticles serving as a receptor is connected with the reporter probes; the capture probes and the reporter probes are in complementary pairing by means of a part of basic groups to form energy transfer donor-and-receptor pairs; the penetrating peptide is connected with the quantum dot to modify the energy transfer donor-and-receptor pairs; the capture probes can be in complementary pairing completely with a target mRNA. The nanoprobe is capable of entering the cells effectively and high in photobleaching resistance. Various nanoprobes are introduced into the living cells to realize real-time parallel detection of content of various mRNA in the living cells; thus, comprehensive understanding on expression levels of different genes in the cells and interaction between the different genes is guaranteed.

Description

technical field [0001] The invention belongs to the technical field of nucleic acid detection, and in particular relates to a nanoprobe for real-time parallel detection of multiple mRNA contents in living cells. Background technique [0002] Genes are the material basis of hereditary variation and dominate the basic structure and performance of life. Gene sequence analysis is the most direct and important part of its research, and it is also the basis for further gene modification. Traditional gene analysis is aimed at population cells, and the result is only the average value of the cell population signal or the information of the dominant number of cells. Even single cells from the same source are different from each other in many aspects due to random biological processes and environmental disturbances, that is, cells have heterogeneity. Compared with the existing gene sequencing technology, single-cell sequencing technology not only helps to understand cell heterogenei...

AI Technical Summary

Problems solved by technology

[0004] In 2005, Zhang et al. from Johns Hopkins University in the United States developed a sandwich-type luminescent probe based on quantum dots and Cy5 dyes, which can be used to determine the content of nucleic acids. Unfortunately, this detection requires complex nucleus extraction, etc. The pretreatment process cannot be applied to the detection of live cells

In 2010, Taejoon et al. of the Korea Institute of Science and Technology used target DNA single strands to connect nano-gold particles and gold nanowires, and observed the Surface Raman Enhanced Effect (SERS), realizing the parallel detection of nucleic acids of various pathogens. However, the material synthesis process of this method is cumbersome, the operation of nanowires is difficult, and the Raman spectroscopy detection equipment is relatively expensive.

In 2014, Michaelis et al. from Germany used the change of the connection position of fluorescent molecules before and after hybridization of nucleic acid molecules to realize the change of fluorescence resonance transfer efficiency, thereby detecting the content of target nucleic acid molecules. Parallel detection of multiple nucleic acids

For example, the expression and transportation of cancer cell growth signals usually do not act alone, but when and in what order do they act. The answers to these questions cannot be given by current detection methods

Some mRNAs are only expressed at a certain stage of the cell physiological cycle, and current detection methods cannot monitor such transient activity changes of mRNAs

Images