Quantitative imaging of single mRNA splice variants in living cells

Abstract

Alternative mRNA splicing is a fundamental process of gene regulation via the precise control of the post-transcriptional step that occurs before mRNA translation. Errors in RNA splicing have been known to correlate with different diseases; however, a key limitation is the lack of technologies for live cell monitoring and quantification to understand the process of alternative splicing. Here, we report a spectroscopic strategy for quantitative imaging of mRNA splice variants in living cells, using nanoplasmonic dimer antennas. The spatial and temporal distribution of three selected splice variants of the breast cancer susceptibility gene, BRCA1 were monitored at single copy resolution by measuring the hybridization dynamics of nanoplasmonic antennas targeting complementary mRNA sequences in live cells. Our study provides valuable insights on RNA and its transport in living cells, which has the potential to enhance our understanding of cellular protein complex, pharmacogenomics, genetic diagnosis, and gene therapies.

Single-cell screening and quantification of transcripts in cancer tissues by second-harmonic generation microscopy

“…4 The nanoprobes are capable of hybridizing to the transcripts with a very broad range of sequences as long as sequence specificity is ensured for transcript targeting. 41 Quantification of mRNA molecules in breast cancer tissues is much lower than that in SK-BR-3 cells, while it is comparable to the number in MCF-7 cells, 30,39 and is in agreement with previous publications. 42 There is large variation in the expression level of Her2 in different cell lines.…”

Graphene laminated gold bipyramids as sensitive detection platforms for antibiotic molecules

“…Localized surface plasmon resonances (LSPR) of plasmonic nanostructures have strong extinction, scattering, and absorption properties are dimension tunable 8–12 and could serve as potential sensor substrates for label-free sensing of a range of targets. It has also been shown that near-field enhancements at the sharp plasmonic edges and strengthening of inter-particle optical coupling, critical for signal enhancement can be observed due to LSPR 13 . LSPR has been exploited to develop highly sensitive biological assays, molecular imaging, photodynamic therapy, drug delivery, and surface-enhanced Raman spectroscopy (SERS).…”

Single-Cell Quantification of Cytosine Modifications by Hyperspectral Dark-Field Imaging

“…35-37 To uncover epigenomic heterogeneity, modern optical microscopy provides elegant solutions for inspecting a single cell. 18, 38, 39 …”

“…18, 19 Owing to the dipole resonance from the interaction with incident photons, the large scattering cross-section of metal NPs can generate a ten- to million-fold stronger signal than conventional fluorophores, 20, 21 providing a high SNR without laser excitation. Besides, the LSPR spectrum can be fine-tuned, dependent on the NP size, shape, material, and surrounding environment.…”