An ultrasensitive bunge bedstraw herb type DNA machine for absolute quantification of mRNA in single cell

Messenger ribonucleic acids (mRNAs) comprise a class of small nucleic acids carrying genetic information, which exhibit very important role in medical research and diagnosis. If only the mean mRNA expression levels of the mRNA population are considered in medical research, important information linking mRNA expression and cellular function may be lost. Single-cell analysis provides valuable insights into studying its heterogeneity, signaling, and stochastic gene expression. In this study, a "bunge bedstraw herb"-type DNA machine based on DNAzyme catalyzing coupled clamping hybrid chain reaction (c-HCR) is presented. In the DNA machine, a bunge bedstraw herb-type DNA structure was first formed by hybridizing a core junction scaffold cruciform probe to a hairpin probe that can trigger the c-HCR via a target molecule in four directions. This approach can reduce the detection limit of mRNA to 5 × 10^-15 M. Absolute quantification of survivin mRNA in individual cells was achieved using the DNA machine on a microfluidic chip electrophoresis platform. The reported method represents an unprecedented single-cell analysis platform for single-cell biology studies.