{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,10,5]],"date-time":"2026-10-05T12:27:24Z","timestamp":1791203244688,"version":"4.2.0"},"posted":{"date-parts":[[2024,10,2]]},"group-title":"Neuroscience","reference-count":105,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2024,10,2]],"date-time":"2024-10-02T00:00:00Z","timestamp":1727827200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2024,12,11]]},"abstract":"<jats:title>SUMMARY<\/jats:title>\n                <jats:p>Single cell genomics has revolutionized our understanding of neuronal cell types. However, scalable technologies for probing single-cell connectivity are lacking, and we are just beginning to understand how molecularly defined cell types are organized into functional circuits. Here, we describe a protocol to generate high-complexity barcoded rabies virus (RV) for scalable circuit mapping from tens of thousands of individual starter cells in parallel. In addition, we introduce a strategy for targeting RV-encoded barcode transcripts to the nucleus so that they can be read out using single-nucleus RNA sequencing (snRNA-seq). We apply this tool in organotypic slice cultures of the developing human cerebral cortex, which reveals the emergence of cell type\u2013 specific circuit motifs in midgestation. By leveraging the power and throughput of single cell genomics for mapping synaptic connectivity, we chart a path forward for scalable circuit mapping of molecularly-defined cell types in healthy and disease states.<\/jats:p>","DOI":"10.1101\/2024.10.01.616167","type":"posted-content","created":{"date-parts":[[2024,10,2]],"date-time":"2024-10-02T09:05:12Z","timestamp":1727859912000},"source":"Crossref","is-referenced-by-count":8,"title":["High-Complexity Barcoded Rabies Virus for Scalable Circuit Mapping Using Single-Cell and Single-Nucleus Sequencing"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-7029-089X","authenticated-orcid":false,"given":"David","family":"Shin","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0320-2260","authenticated-orcid":false,"given":"Madeleine E.","family":"Urbanek","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"H. Hanh","family":"Larson","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9538-2747","authenticated-orcid":false,"given":"Anthony J.","family":"Moussa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kevin Y.","family":"Lee","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Donovan L.","family":"Baker","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Elio","family":"Standen-Bloom","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-9674-3310","authenticated-orcid":false,"given":"Sangeetha","family":"Ramachandran","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Derek","family":"Bogdanoff","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1963-8285","authenticated-orcid":false,"given":"Cathryn R.","family":"Cadwell","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tomasz J.","family":"Nowakowski","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2024121402253677000_2024.10.01.616167v2.1","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth999"},{"key":"2024121402253677000_2024.10.01.616167v2.2","doi-asserted-by":"publisher","DOI":"10.1038\/nature09714"},{"key":"2024121402253677000_2024.10.01.616167v2.3","doi-asserted-by":"publisher","DOI":"10.1126\/science.abf1230"},{"key":"2024121402253677000_2024.10.01.616167v2.4","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-022-34334-1"},{"key":"2024121402253677000_2024.10.01.616167v2.5","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.87866"},{"key":"2024121402253677000_2024.10.01.616167v2.6","doi-asserted-by":"publisher","DOI":"10.1038\/nprot.2009.248"},{"key":"2024121402253677000_2024.10.01.616167v2.7","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.73.5.3818-3825.1999"},{"key":"2024121402253677000_2024.10.01.616167v2.8","doi-asserted-by":"publisher","DOI":"10.1002\/j.1460-2075.1994.tb06739.x"},{"key":"2024121402253677000_2024.10.01.616167v2.9","unstructured":"Negri, A. , and Foundation, C.S. (1905). Contributo allo studio dell\u2019eziologia della rabbia (Tipografia e Legatoria Cooperativa)."},{"key":"2024121402253677000_2024.10.01.616167v2.10","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.00554-09"},{"key":"2024121402253677000_2024.10.01.616167v2.11","doi-asserted-by":"publisher","DOI":"10.1038\/S41586-021-04230-7"},{"key":"2024121402253677000_2024.10.01.616167v2.12","doi-asserted-by":"publisher","DOI":"10.1145\/584091.584093"},{"key":"2024121402253677000_2024.10.01.616167v2.13","doi-asserted-by":"publisher","DOI":"10.1016\/j.plrev.2004.01.002"},{"key":"2024121402253677000_2024.10.01.616167v2.14","doi-asserted-by":"publisher","DOI":"10.1101\/pdb.prot072389"},{"key":"2024121402253677000_2024.10.01.616167v2.15","doi-asserted-by":"publisher","DOI":"10.1038\/nprot.2013.094"},{"key":"2024121402253677000_2024.10.01.616167v2.16","doi-asserted-by":"publisher","DOI":"10.1016\/j.virusres.2015.05.023"},{"key":"2024121402253677000_2024.10.01.616167v2.17","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.79848"},{"key":"2024121402253677000_2024.10.01.616167v2.18","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2016.03.067"},{"key":"2024121402253677000_2024.10.01.616167v2.19","unstructured":"Ian Wickersham , Shenqin Yao , Bosiljka Tasic , Xiaoyin Chen (2024). Highly multiplexed circuit mapping using barcoded rabies viruses and in situ sequencing."},{"key":"2024121402253677000_2024.10.01.616167v2.20","doi-asserted-by":"publisher","DOI":"10.1016\/0006-8993(73)90741-5"},{"key":"2024121402253677000_2024.10.01.616167v2.21","doi-asserted-by":"publisher","DOI":"10.1002\/cne.902970309"},{"key":"2024121402253677000_2024.10.01.616167v2.22","doi-asserted-by":"publisher","DOI":"10.1038\/s41592-019-0619-0"},{"key":"2024121402253677000_2024.10.01.616167v2.23","doi-asserted-by":"publisher","DOI":"10.1038\/nbt.4314"},{"key":"2024121402253677000_2024.10.01.616167v2.24","doi-asserted-by":"publisher","DOI":"10.1088\/1742-5468\/2008\/10\/P10008"},{"key":"2024121402253677000_2024.10.01.616167v2.25","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2008.08.006"},{"key":"2024121402253677000_2024.10.01.616167v2.26","doi-asserted-by":"publisher","DOI":"10.1016\/j.stem.2018.05.024"},{"key":"2024121402253677000_2024.10.01.616167v2.27","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2010.04.038"},{"key":"2024121402253677000_2024.10.01.616167v2.28","doi-asserted-by":"publisher","DOI":"10.1016\/j.brainres.2018.02.044"},{"key":"2024121402253677000_2024.10.01.616167v2.29","doi-asserted-by":"publisher","DOI":"10.1016\/j.tins.2010.10.002"},{"key":"2024121402253677000_2024.10.01.616167v2.30","doi-asserted-by":"publisher","DOI":"10.1101\/2024.06.05.597673"},{"key":"2024121402253677000_2024.10.01.616167v2.31","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2012.02.052"},{"key":"2024121402253677000_2024.10.01.616167v2.32","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.52370"},{"key":"2024121402253677000_2024.10.01.616167v2.33","first-page":"687","article-title":"A comparison of Notch, Hes and Grg expression during murine embryonic and post-natal development","volume":"45","year":"1999","journal-title":"Cell. Mol. Biol"},{"key":"2024121402253677000_2024.10.01.616167v2.34","doi-asserted-by":"publisher","DOI":"10.1038\/nrn2151"},{"key":"2024121402253677000_2024.10.01.616167v2.35","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2016.05.024"},{"key":"2024121402253677000_2024.10.01.616167v2.36","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2023.112957"},{"key":"2024121402253677000_2024.10.01.616167v2.37","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2004.12.036"},{"key":"2024121402253677000_2024.10.01.616167v2.38","doi-asserted-by":"publisher","DOI":"10.1016\/J.CELL.2021.07.039"},{"key":"2024121402253677000_2024.10.01.616167v2.39","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-021-03209-8"},{"key":"2024121402253677000_2024.10.01.616167v2.40","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.5963-09.2010"},{"key":"2024121402253677000_2024.10.01.616167v2.41","doi-asserted-by":"publisher","DOI":"10.1093\/cercor\/bhk003"},{"key":"2024121402253677000_2024.10.01.616167v2.42","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-023-06812-z"},{"key":"2024121402253677000_2024.10.01.616167v2.43","doi-asserted-by":"publisher","DOI":"10.1242\/dev.117.2.525"},{"key":"2024121402253677000_2024.10.01.616167v2.44","doi-asserted-by":"publisher","DOI":"10.1242\/dev.115.2.535"},{"key":"2024121402253677000_2024.10.01.616167v2.45","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.2899-04.2005"},{"key":"2024121402253677000_2024.10.01.616167v2.46","doi-asserted-by":"publisher","DOI":"10.1242\/dev.022616"},{"key":"2024121402253677000_2024.10.01.616167v2.47","doi-asserted-by":"publisher","DOI":"10.1016\/j.neures.2006.03.004"},{"key":"2024121402253677000_2024.10.01.616167v2.48","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2015.09.004"},{"key":"2024121402253677000_2024.10.01.616167v2.49","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.20-01-00283.2000"},{"key":"2024121402253677000_2024.10.01.616167v2.50","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M102420200"},{"key":"2024121402253677000_2024.10.01.616167v2.51","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbamem.2006.02.018"},{"key":"2024121402253677000_2024.10.01.616167v2.52","doi-asserted-by":"publisher","DOI":"10.1152\/physrev.00011.2013"},{"key":"2024121402253677000_2024.10.01.616167v2.53","doi-asserted-by":"publisher","DOI":"10.1016\/j.neulet.2014.10.040"},{"key":"2024121402253677000_2024.10.01.616167v2.54","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.14-03-01030.1994"},{"key":"2024121402253677000_2024.10.01.616167v2.55","doi-asserted-by":"publisher","DOI":"10.1111\/joa.12948"},{"key":"2024121402253677000_2024.10.01.616167v2.56","doi-asserted-by":"publisher","DOI":"10.1007\/s11064-004-6881-1"},{"key":"2024121402253677000_2024.10.01.616167v2.57","doi-asserted-by":"publisher","DOI":"10.1002\/ijc.2910310104"},{"key":"2024121402253677000_2024.10.01.616167v2.58","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2016.01.004"},{"key":"2024121402253677000_2024.10.01.616167v2.59","doi-asserted-by":"publisher","DOI":"10.1038\/sj.gt.3301905"},{"key":"2024121402253677000_2024.10.01.616167v2.60","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2010.08.001"},{"key":"2024121402253677000_2024.10.01.616167v2.61","doi-asserted-by":"publisher","DOI":"10.3389\/fncel.2021.720807"},{"key":"2024121402253677000_2024.10.01.616167v2.62","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.02044-14"},{"key":"2024121402253677000_2024.10.01.616167v2.63","first-page":"2031","article-title":"Abortively infected astrocytes appear to represent the main source of interferon beta in the virus-infected brain","volume":"90","year":"2015","journal-title":"J. Virol"},{"key":"2024121402253677000_2024.10.01.616167v2.64","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1618157114"},{"key":"2024121402253677000_2024.10.01.616167v2.65","doi-asserted-by":"publisher","DOI":"10.1371\/JOURNAL.PONE.0209648"},{"key":"2024121402253677000_2024.10.01.616167v2.66","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.2203677119"},{"key":"2024121402253677000_2024.10.01.616167v2.67","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.69.10.3033"},{"key":"2024121402253677000_2024.10.01.616167v2.68","doi-asserted-by":"publisher","DOI":"10.1016\/j.cub.2006.03.092"},{"key":"2024121402253677000_2024.10.01.616167v2.69","doi-asserted-by":"publisher","DOI":"10.1242\/jcs.057075"},{"key":"2024121402253677000_2024.10.01.616167v2.70","doi-asserted-by":"publisher","DOI":"10.1186\/s13059-020-1950-6"},{"key":"2024121402253677000_2024.10.01.616167v2.71","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2024.09.003"},{"key":"2024121402253677000_2024.10.01.616167v2.72","doi-asserted-by":"publisher","DOI":"10.1038\/s41592-018-0185-x"},{"key":"2024121402253677000_2024.10.01.616167v2.73","first-page":"1827","article-title":"A rabies virus-based toolkit for efficient retrograde labeling and monosynaptic tracing","volume":"18","year":"2023","journal-title":"Neural Regeneration Res"},{"key":"2024121402253677000_2024.10.01.616167v2.74","doi-asserted-by":"publisher","DOI":"10.3389\/fbioe.2021.701031"},{"key":"2024121402253677000_2024.10.01.616167v2.75","doi-asserted-by":"publisher","DOI":"10.1016\/j.neuron.2013.08.027"},{"key":"2024121402253677000_2024.10.01.616167v2.76","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-021-98657-7"},{"key":"2024121402253677000_2024.10.01.616167v2.77","doi-asserted-by":"publisher","DOI":"10.1101\/2024.04.04.588058"},{"key":"2024121402253677000_2024.10.01.616167v2.78","doi-asserted-by":"publisher","DOI":"10.3389\/fnana.2019.00041"},{"key":"2024121402253677000_2024.10.01.616167v2.79","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.5169-06.2007"},{"key":"2024121402253677000_2024.10.01.616167v2.80","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.5599-07.2008"},{"key":"2024121402253677000_2024.10.01.616167v2.81","doi-asserted-by":"publisher","DOI":"10.1038\/nature04404"},{"key":"2024121402253677000_2024.10.01.616167v2.82","doi-asserted-by":"publisher","DOI":"10.1016\/j.pneurobio.2020.101883"},{"key":"2024121402253677000_2024.10.01.616167v2.83","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.4748-11.2012"},{"key":"2024121402253677000_2024.10.01.616167v2.84","doi-asserted-by":"publisher","DOI":"10.1093\/cercor\/bhw271"},{"key":"2024121402253677000_2024.10.01.616167v2.85","doi-asserted-by":"publisher","DOI":"10.7554\/elife.100880.1"},{"key":"2024121402253677000_2024.10.01.616167v2.86","doi-asserted-by":"publisher","DOI":"10.1101\/2022.06.22.497212"},{"key":"2024121402253677000_2024.10.01.616167v2.87","doi-asserted-by":"publisher","DOI":"10.1038\/s41592-021-01126-2"},{"key":"2024121402253677000_2024.10.01.616167v2.88","doi-asserted-by":"publisher","DOI":"10.2144\/0000113878"},{"key":"2024121402253677000_2024.10.01.616167v2.89","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.4407"},{"key":"2024121402253677000_2024.10.01.616167v2.90","doi-asserted-by":"crossref","unstructured":"Martelotto, L.G. , Kalavros, N. , Vlachos, I. , and Wang, S. (2023). BIDMC TMC - low input single nuclei sequencing isolation (NNLB).","DOI":"10.17504\/protocols.io.x54v9pp8qg3e\/v1"},{"key":"2024121402253677000_2024.10.01.616167v2.91","unstructured":"Bushnell, B. (2022). BBMap. SourceForge. https:\/\/sourceforge.net\/projects\/bbmap\/."},{"key":"2024121402253677000_2024.10.01.616167v2.92","doi-asserted-by":"publisher","DOI":"10.1101\/gr.209601.116"},{"key":"2024121402253677000_2024.10.01.616167v2.93","doi-asserted-by":"publisher","DOI":"10.1109\/MCSE.2007.55"},{"key":"2024121402253677000_2024.10.01.616167v2.94","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-020-2649-2"},{"key":"2024121402253677000_2024.10.01.616167v2.95","doi-asserted-by":"publisher","DOI":"10.5281\/zenodo.3509134"},{"key":"2024121402253677000_2024.10.01.616167v2.96","doi-asserted-by":"publisher","DOI":"10.21105\/joss.03021"},{"key":"2024121402253677000_2024.10.01.616167v2.97","doi-asserted-by":"publisher","DOI":"10.1016\/J.CELL.2021.04.048"},{"key":"2024121402253677000_2024.10.01.616167v2.98","doi-asserted-by":"crossref","unstructured":"Korsunsky, I. , Nathan, A. , Millard, N. , and Raychaudhuri, S . (2019). Presto scales Wilcoxon and auROC analyses to millions of observations. Bioinformatics.","DOI":"10.1101\/653253"},{"key":"2024121402253677000_2024.10.01.616167v2.99","unstructured":"Blighe, K. , Rana, S. , Lewis, M. EnhancedVolcano: Publication-ready volcano plots with enhanced colouring and labeling (Github)."},{"key":"2024121402253677000_2024.10.01.616167v2.100","doi-asserted-by":"publisher","DOI":"10.1089\/omi.2011.0118"},{"key":"2024121402253677000_2024.10.01.616167v2.101","first-page":"100141","article-title":"clusterProfiler 4.0: A universal enrichment tool for interpreting omics data","volume":"2","year":"2021","journal-title":"Innovation (Camb"},{"key":"2024121402253677000_2024.10.01.616167v2.102","doi-asserted-by":"publisher","DOI":"10.1016\/j.csbj.2021.06.043"},{"key":"2024121402253677000_2024.10.01.616167v2.103","doi-asserted-by":"publisher","DOI":"10.1038\/s41592-019-0686-2"},{"key":"2024121402253677000_2024.10.01.616167v2.104","doi-asserted-by":"crossref","unstructured":"Aric, A. , Hagberg, D.A. , and Schult, P.J. (2008). Exploring network structure, dynamics, and function using NetworkX\u201d. In Proceedings of the 7th Python in Science Conference (SciPy2008), G\u00e4el Varoquaux, T. Vaught and J. Millman, eds., pp. 11\u201315.","DOI":"10.25080\/TCWV9851"},{"key":"2024121402253677000_2024.10.01.616167v2.105","doi-asserted-by":"publisher","DOI":"10.1002\/1097-024X(200009)30:11<1203::AID-SPE338>3.0.CO;2-N"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1101\/2024.10.01.616167","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T07:11:13Z","timestamp":1768461073000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.1101\/2024.10.01.616167"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,10,2]]},"references-count":105,"URL":"https:\/\/doi.org\/10.1101\/2024.10.01.616167","relation":{},"subject":[],"published":{"date-parts":[[2024,10,2]]},"subtype":"preprint"}}