{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,9,27]],"date-time":"2026-09-27T14:57:49Z","timestamp":1790521069961,"version":"4.1.0"},"posted":{"date-parts":[[2026,9,22]]},"group-title":"Developmental Biology","reference-count":47,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,9,22]],"date-time":"2026-09-22T00:00:00Z","timestamp":1790035200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nd\/4.0\/"}],"funder":[{"name":"Eunice Kennedy Shriver National Institute of Child Health and Human Development","award":["1R21HD119496-01"],"award-info":[{"award-number":["1R21HD119496-01"]}],"id":[{"id":"https:\/\/ror.org\/04byxyr05","id-type":"ROR","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,9,22]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Regenerating tissues like the axolotl (\n                  <jats:italic>Ambystoma mexicanum<\/jats:italic>\n                  ) limb draw from many cell types, but isolating specific cell populations remains a bottleneck. Standard approaches rely on enzymatic dissociation and fluorescence-activated cell sorting (FACS), which are slow, costly, and require specialized instrumentation. We adapted INTACT (Isolation of\n                  <jats:underline>N<\/jats:underline>\n                  uclei\n                  <jats:underline>Ta<\/jats:underline>\n                  gged in specific\n                  <jats:underline>C<\/jats:underline>\n                  ell\n                  <jats:underline>T<\/jats:underline>\n                  ypes) to the axolotl, generating a transgenic line (\n                  <jats:italic>INTACT-olotl<\/jats:italic>\n                  ) expressing\n                  <jats:italic>Sun1-sfGFP<\/jats:italic>\n                  in limb connective tissue, the population that carries limb positional information, dedifferentiates following injury, and forms the majority of the regenerative blastema. Combining an optimized nuclei isolation protocol with anti-GFP nanobody capture enriched labeled nuclei from \u223c34.6% of input to 80\u201389% purity. Relative to dissociation and FACS, this recovers roughly 30-fold more target nuclei in less than half the time and without a cytometer. Bead-bound nuclei are directly applied to combinatorial-barcoding snRNA-seq, eliminating fluorescence-activated nuclei sorting (FANS). Profiling INTACT\n                  <jats:sup>+<\/jats:sup>\n                  connective tissue during TGF\u03b2 inhibition revealed a loss of extracellular matrix remodeling with upregulated pro-fibrotic and inflammatory gene programs, resembling an unresolved wound environment. These changes reverse and regeneration resumes upon drug washout, indicating that functional TGF\u03b2 signaling is required for the transition from wound resolution to blastema formation.\n                <\/jats:p>","DOI":"10.64898\/2026.09.18.752769","type":"posted-content","created":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T19:00:25Z","timestamp":1790362825000},"source":"Crossref","is-referenced-by-count":0,"title":["<i>INTACT-olotl<\/i>\n                  : A simplified system for rapid cell type-specific profiling during axolotl limb regeneration"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0009-0007-1507-3201","authenticated-orcid":false,"given":"Jacob E.","family":"Barrett","sequence":"first","affiliation":[{"name":"Wake Forest University, Department of Biology, Winston-Salem, NC"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ramya","family":"Ailuri","sequence":"additional","affiliation":[{"name":"Wake Forest University, Department of Biology, Winston-Salem, NC"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zinn","family":"Amos","sequence":"additional","affiliation":[{"name":"Wake Forest University, Department of Biology, Winston-Salem, NC"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tim","family":"Curtis","sequence":"additional","affiliation":[{"name":"Wake Forest University, Department of Biology, Winston-Salem, NC"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Samantha","family":"Rock","sequence":"additional","affiliation":[{"name":"Wake Forest University, Department of Biology, Winston-Salem, NC"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-4871-0525","authenticated-orcid":false,"given":"Joshua D.","family":"Currie","sequence":"additional","affiliation":[{"name":"Wake Forest University, Department of Biology, Winston-Salem, NC"},{"name":"Wake Forest Institute of Regenerative Medicine; Winston-Salem, NC, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026092707464120000_2026.09.18.752769v1.1","doi-asserted-by":"crossref","unstructured":"Ahuja, G. , Antill, A. , Su, Y. , Dall\u2019Olio, G. M. , Basnayake, S. , Karlsson, G. and Dhapola, P. (2025). 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Target. Ther"},{"key":"2026092707464120000_2026.09.18.752769v1.9","doi-asserted-by":"publisher","DOI":"10.1016\/S0014-4827(02)00015-0"},{"key":"2026092707464120000_2026.09.18.752769v1.10","doi-asserted-by":"crossref","first-page":"195","DOI":"10.1089\/wound.2013.0429","article-title":"Dynamics of transforming growth factor beta signaling in wound healing and scarring","volume":"2","year":"2013","journal-title":"Adv. Wound Care (New Rochelle"},{"key":"2026092707464120000_2026.09.18.752769v1.11","doi-asserted-by":"publisher","DOI":"10.1007\/s10555-020-09909-3"},{"key":"2026092707464120000_2026.09.18.752769v1.12","doi-asserted-by":"crossref","unstructured":"Garc\u00eda-Garc\u00eda, D. , Knapp, D. , Kim, M. , Jamwal, K. , Fuqua, H. , Seaman, R. P. , Grindle, R. E. , Nowoshilow, S. , Novatchkova, M. , Kolling, F. W. , et al. (2025). 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