Quantitative Super-Resolution Imaging of Small RNAs in Bacterial Cells
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We present a method for the quantification of small regulatory RNAs (sRNAs) in bacteria, by combining single-molecule fluorescence in situ hybridization (smFISH), super-resolved single-fluorophore microscopy, and clustering analysis. Compared to smFISH imaging with diffraction-limited fluorescence microscopy, our method provides better quantification for short and abundant RNA (such as sRNAs) in a small volume of bacterial cells. Our method can also be directly used for the quantification of messenger RNAs (mRNAs).
KeywordsFluorescence in situ hybridization Super-resolution microscopy Clustering analysis Small regulatory RNA
The data presented as examples were collected during the 2016 Center for the Physics of Living Cells (CPLC) Summer School at University of Illinois at Urbana-Champaign. We therefore thank National Science Foundation grant PHY-1430124 (Physics Frontiers Center for the Physics of Living Cells) for funding for the CPLC Summer School, Zan Luthey-Schulten and Joseph R. Peterson for co-teaching the scientific theme of “Quantitative Imaging and Cell Simulation of Small Regulatory RNA,” and the summer school students: Bijoy Desai (Columbia University), Dennis Fernandes (University of Toronto, Mississauga), Jialei Tang (University of Central Florida), and Kailun Zhang (Texas A&M University) for participation in data collection and analysis.