Ninu (greater bilby, Macrotis lagotis) are desert-dwelling, culturally and ecologically important marsupials. In collaboration with Indigenous rangers and conservation managers, we generated the Ninu chromosome-level genome assembly (3.66 Gbp) and genome sequences for the extinct Yallara (lesser bilby, Macrotis leucura). We developed and tested a scat single-nucleotide polymorphism panel to inform current and future conservation actions, undertake ecological assessments and improve our understanding of Ninu genetic diversity in managed and wild populations. We also assessed the beneficial impact of translocations in the metapopulation (N = 363 Ninu). Resequenced genomes (temperate Ninu, 6; semi-arid Ninu, 6; and Yallara, 4) revealed two major population crashes during global cooling events for both species and differences in Ninu genes involved in anatomical and metabolic pathways. Despite their 45-year captive history, Ninu have fewer long runs of homozygosity than other larger mammals, which may be attributable to their boom–bust life history. Here we investigated the unique Ninu biology using 12 tissue transcriptomes revealing expression of all 115 conserved eutherian chorioallantoic placentation genes in the uterus, an XY₁Y₂ sex chromosome system and olfactory receptor gene expansions. Together, we demonstrate the holistic value of genomics in improving key conservation actions, understanding unique biological traits and developing tools for Indigenous rangers to monitor remote wild populations.

尼努（大兔耳袋狸，Macrotis lagotis）是生活在沙漠中、具有重要文化和生态意义的有袋动物。我们与土著护林员和保护管理者合作，生成了 Ninu 染色体水平基因组组装 (3.66 Gbp) 和已灭绝的 Yallara（小兔耳袋狸，Macrotis leucura）的基因组序列。我们开发并测试了粪便单核苷酸多态性面板，为当前和未来的保护行动提供信息，进行生态评估并提高我们对管理和野生种群中尼努遗传多样性的了解。我们还评估了易位对集合种群的有益影响（N = 363 Ninu）。基因组重新测序（温带 Ninu，6；半干旱 Ninu，6；和 Yallara，4）揭示了这两个物种在全球变冷事件期间发生的两次主要种群崩溃，以及涉及解剖和代谢途径的 Ninu 基因的差异。尽管有 45 年的圈养历史，尼努的长期纯合性比其他大型哺乳动物要少，这可能归因于它们的繁荣-萧条生活史。在这里，我们使用 12 个组织转录组研究了独特的 Ninu 生物学，揭示了子宫中所有 115 个保守的真兽类绒毛膜尿囊胎盘基因的表达、XY ₁ Y ₂ 性染色体系统和嗅觉受体基因扩展。我们共同证明了基因组学在改善关键保护行动、了解独特的生物特征以及为土著护林员开发监测偏远野生种群的工具方面的整体价值。