@article{f1cfff4cb1b648e8bc5e5508290d3a69,
title = "Polony gels enable amplifiable DNA stamping and spatial transcriptomics of chronic pain",
abstract = "Methods for acquiring spatially resolved omics data from complex tissues use barcoded DNA arrays of low- to sub-micrometer features to achieve single-cell resolution. However, fabricating such arrays (randomly assembled beads, DNA nanoballs, or clusters) requires sequencing barcodes in each array, limiting cost-effectiveness and throughput. Here, we describe a vastly scalable stamping method to fabricate polony gels, arrays of ∼1-micrometer clonal DNA clusters bearing unique barcodes. By enabling repeatable enzymatic replication of barcode-patterned gels, this method, compared with the sequencing-dependent array fabrication, reduced cost by at least 35-fold and time to approximately 7 h. The gel stamping was implemented with a simple robotic arm and off-the-shelf reagents. We leveraged the resolution and RNA capture efficiency of polony gels to develop Pixel-seq, a single-cell spatial transcriptomic assay, and applied it to map the mouse parabrachial nucleus and analyze changes in neuropathic pain-regulated transcriptomes and cell-cell communication after nerve ligation.",
keywords = "DNA array, DNA stamping, Pixel-seq, chronic pain, microcontact printing, olfactory bulb, parabrachial nucleus, polony gel, polony sequencing, spatial transcriptomics",
author = "Xiaonan Fu and Li Sun and Runze Dong and Chen, {Jane Y.} and Runglawan Silakit and Condon, {Logan F.} and Yiing Lin and Shin Lin and Palmiter, {Richard D.} and Liangcai Gu",
note = "Funding Information: We thank B. Wang and W. Liang for contributions to a base calling algorithm, D. Liu and J. Cheng for the help to build the polony sequencer, Q. Tang for sharing mouse tissues, and S. Kang for discussing assay development. This work was supported by a University of Washington startup fund to L.G. and National Institutes of Health grants (R35GM128918, UG3CA268096, R21DA051555, and R21DA051194 to L.G. R41MH130299 to L.S. and R01DA024908 to R.D.P.). L.G. conceived the methodology and supervised this project. L.S. and L.G. developed the polony stamping. X.F. L.G. and L.S. developed the Pixel-seq assay. L.S. and R.S. fabricated polony gels. X.F. R.D. L.G. S.Y. and Y.L. analyzed the data. J.Y.C. L.F.C. and R.D.P. designed the PBN study and prepared the tissues. L.G. X.F. and R.D. wrote the manuscript with input from other co-authors. Two patents on polony gel stamping and Pixel-seq have been filed by the University of Washington and TopoGene Inc. L.G. and L.S. are co-founders of TopoGene, which aims to commercialize polony gels. Funding Information: We thank B. Wang and W. Liang for contributions to a base calling algorithm, D. Liu and J. Cheng for the help to build the polony sequencer, Q. Tang for sharing mouse tissues, and S. Kang for discussing assay development. This work was supported by a University of Washington startup fund to L.G. and National Institutes of Health grants (R35GM128918, UG3CA268096, R21DA051555, and R21DA051194 to L.G., R41MH130299 to L.S., and R01DA024908 to R.D.P.). Publisher Copyright: {\textcopyright} 2022 Elsevier Inc.",
year = "2022",
month = nov,
day = "23",
doi = "10.1016/j.cell.2022.10.021",
language = "English",
volume = "185",
pages = "4621--4633.e17",
journal = "Cell",
issn = "0092-8674",
number = "24",
}