Beyond quantification: in situ analysis of transcriptome and pre-mRNA alternative splicing at the nanoscale.
In situ analysis offers a venue for dissecting the complex transcriptome in its natural context to tap into cellular processes that could explain the phenotypic physiology and pathology yet to be understood. Over the past decades, enormous progress has been made to improve the resolution, sensitivity, and specificity of single-cell technologies. The continued efforts in RNA research not only facilitates mechanistic studies of molecular biology but also provides state-of-the-art strategies for diagnostic purposes. The implementation of novel bio-imaging platforms has yielded valuable information for inspecting gene expression, mapping regulatory networks, and classifying cell types. In this article, we discuss the merits and technical challenges in single-molecule in situ RNA profiling. Advanced in situ hybridization methodologies developed for a variety of detection modalities are reviewed. Considering the fact that in mammalian cells the number of protein products immensely exceeds that of the actual coding genes due to pre-mRNA alternative splicing, tools capable of elucidating this process in intact cells are highlighted. To conclude, we point out future directions for in situ transcriptome analysis and expect a plethora of opportunities and discoveries in this field. WIREs Nanomed Nanobiotechnol 2017, 9:e1443. doi: 10.1002/wnan.1443 For further resources related to this article, please visit the WIREs website.
Department of Agricultural and Biological Engineering, Bindley Bioscience Center and Birck Nanotechnology Center, Purdue Center for Cancer Research, Purdue University, West Lafayette, IN, USA.|Environmental Molecular Science Laboratory, Pacific Northwest National Laboratory, Richland, WA, USA.
Yi
Y
Cui
Department of Nanoscience and Nanoengineering, South Dakota School of Mines & Technology, Rapid City, SD, USA.
Jing
J
Liu
Department of Agricultural and Biological Engineering, Bindley Bioscience Center and Birck Nanotechnology Center, Purdue Center for Cancer Research, Purdue University, West Lafayette, IN, USA.
Joseph
J
Irudayaraj
D017398:Alternative Splicing; D000818:Animals; D020869:Gene Expression Profiling; D006801:Humans; D017403:In Situ Hybridization; D008853:Microscopy; D053758:Nanoparticles; D012322:RNA Precursors / Q000235:genetics*; D059467:Transcriptome
D012322:RNA Precursors