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Running title: Ontogeny of burrowing in deer mice SummaryA central challenge in biology is to understand how innate behaviors evolve between closely related species. One way to elucidate how differences arise is to compare the development of behavior in species with distinct adult traits. Here, we report that Peromyscus polionotus is strikingly precocious with regard to burrowing behavior, but not other behaviors, compared to its sister species P. maniculatus. In P. polionotus, burrows were excavated as early as 17 days of age, while P. maniculatus did not build burrows until 10 days later. Moreover, the well-known differences in burrow architecture between adults of these species-P. polionotus adults excavate long burrows with an escape tunnel, while P. maniculatus dig short, single-tunnel burrows-were intact in juvenile burrowers. To test whether this juvenile behavior is influenced by early-life environment, pups of both species were reciprocally cross-fostered. Fostering did not alter the characteristic burrowing behavior of either species, suggesting these differences are genetic. In backcross F2 hybrids, we show that precocious burrowing and adult tunnel length are genetically correlated, and that a single P. polionotus allele in a genomic region linked to adult tunnel length is predictive of precocious burrow construction. The co-inheritance of developmental and adult traits indicates the same genetic region-either a single gene with pleiotropic effects, or closely linked genesacts on distinct aspects of the same behavior across life stages. Such genetic variants likely affect behavioral drive (i.e. motivation) to burrow, and thereby affect both the development and adult expression of burrowing behavior. The copyright holder for this preprint (which was not . http://dx.doi.org/10.1101/150243 doi: bioRxiv preprint first posted online Jun. 14, 2017; 2 Highlights  Juvenile P. polionotus construct burrows precociously compared to its sister species P. maniculatus  Cross-fostering does not alter species-specific burrowing behavior  A QTL linked to adult tunnel length predicts developmental onset of burrow construction in hybrids  Pleiotropic genetic variant(s) may affect behavioral drive across life stages eTOC Metz et al. find that oldfield mice, a species that digs long, complex burrows, also digs burrows earlier in development compared to its sister species. In F2 hybrids, precocious burrowing is co-inherited with long adult tunnels, and an allele linked to adult tunnel length also predicts timing of first burrow construction, suggesting that a single genetic region controls different aspects of the same behavior across distinct life stages.peer-reviewed) is the author/funder. All rights reserved. No reuse allowed without permission.The copyright holder for this preprint (which was not . http://dx.doi.org/10.1101/150243 doi: bioRxiv preprint first posted online Jun. 14, 2017; 3 Results P. POLIONOTUS CONSTRUCT BURROWS EARLIER IN LIFE THAN P. MANICULATUSTo examine the developmental onset of burrow cons...