EDGE FGT: Functional Genomics in Gar - Discovery Tools for Major Vertebrate Transitions

脊椎动物的生物多样性包括3万种陆地脊椎动物（包括人类在内的四足动物）和3万种现代鱼类（硬骨鱼）。直接比较这两个群体仍然具有挑战性，因为它们与4.5亿年前的共同祖先相比发生了重大变化：四足动物的身体在从鱼类出现并开始在陆地上生活时经历了重大转变；硬鱼彻底改变了它们的基因组结构，获得了数千个额外的基因。这个NSF EDGE项目开发了古老而独特的信息garfish （Lepisosteidae）作为研究生物，连接了四足动物和硬骨鱼的不同世界。它们的基因组在组织上与四足动物的基因组相似，而鱼的发育与硬骨鱼非常相似，这使得鱼成为理解脊椎动物进化和我们自己的生物起源的“罗塞塔石碑”。该项目将改进gar养殖和产卵技术，并开发新的基因操作方法，如CRISPR基因组编辑，以测试gar基因功能。这将使gar作为实验物种，在脊椎动物生物学家的广泛社区中进行比较基因组，进化，发育，生理，行为，再生和生态研究。该项目将通过对K-12和本科生的实验室体验、gar会议和脊椎动物进化遗传学研讨会来培养下一代脊椎动物生物学家。公众研讨会、向水族馆和动物园捐赠动物以及强大的社交媒体将通过强调鱼类对理解进化、遗传和发展的重要性，加强公众的科学素养。因此，该项目将提高人们对gar数量下降的生态意义的认识。脊椎动物的进化伴随着特定谱系的基因和形态结构的获得和损失。因此，研究具有信息基因组、发育和表型特征的生物体对于了解脊椎动物的起源、主要转变、生物多样性和基因组功能至关重要。该项目将通过开发斑点雀（Lepisosteus oculatus）的基因组-表型技术来研究脊椎动物进化中的广泛生物学问题，斑点雀（Lepisosteus oculatus）在骨性脊椎动物中占据关键的系统发育位置。gar既是硬骨鱼类在进化早期经历了整个基因组复制事件的实验可获得的“非复制”外群，也是包括四足动物在内的叶鳍脊椎动物的外群。gar基因组组装已在300多个基因组研究中使用，但目前缺乏能够直接检测gar基因型-表型关系的功能遗传学研究。该项目利用正在进行的研究工作，以促进gar的基因组实现。需要开发的资源和遗传工具包括：1)改进鱼卵产卵技术和基础设施，以进行鱼卵分子遗传实验；2)针对gar模型的CRISPR基因组编辑、靶向基因沉默和转基因技术进展；3)通过项目特定的在线数据库、两次Gar会议和两次关于功能脊椎动物Evo-Devo基因组学的研讨会传播项目结果。因此，该项目旨在了解一组gar基因和调控元件的作用，作为原理证明，以解锁gar作为模式生物的全部潜力，用于比较脊椎动物生物学的广泛研究问题。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

The biodiversity of vertebrate animals includes 30,000 land vertebrates (tetrapods, including humans) as well as 30,000 species of modern fishes (teleosts). It remains challenging to compare the two groups directly because they significantly changed from their common ancestor 450 million years ago: tetrapod bodies underwent major transitions when they emerged from fishes and began to live on land; teleosts drastically changed their genome organization and gained thousands of extra genes. This NSF EDGE project develops the archaic and uniquely informative garfishes (Lepisosteidae) as research organisms that bridge the disparate worlds of tetrapods and teleosts. Their genome resembles the tetrapod genome in its organization while gars develop very similar to teleosts, making gars a “Rosetta Stone” to understand vertebrate evolution and our own biological origins. The project will improve gar husbandry and spawning techniques and develop novel methodologies for genetic manipulations such as CRISPR genome editing to test gar gene functions. This will make gars available as experimental species for comparative genomic, evolutionary, developmental, physiological, behavioral, regeneration, and ecological research among the broad community of vertebrate biologists. The project will train the next generation of vertebrate biologists through laboratory experiences for K-12 and undergraduate students, gar conferences, and workshops on vertebrate evolutionary genetics. Public seminars, animal donations to aquaria and zoos, and strong social media presence will strengthen scientific literacy of the general public by emphasizing the importance of fishes for understanding evolution, genetics, and development. Thereby, the project will raise awareness for the ecological significance of declining gar populations.The evolution of vertebrates has been accompanied by lineage-specific gains and losses of genes and morphological structures. Research organisms with informative genomes, development, and phenotypic traits are thus essential to understand vertebrate origins, major transitions, their biodiversity, and genome functions. This project will enable the investigation of broad biological questions across vertebrate evolution by developing genome-to-phenome technologies for the spotted gar (Lepisosteus oculatus) that occupies a key phylogenetic position among bony vertebrates. Gars serve both as an experimentally accessible, "unduplicated" outgroup to the teleost fishes that went through a whole genome duplication event early in their evolution as well as an outgroup to lobe-finned vertebrates including tetrapods. The gar genome assembly has been used in 300+ genomic studies, but functional genetic investigations that enable direct testing of genotype-to-phenotype relations in gar are currently lacking. This project leverages ongoing research efforts in order to facilitate the genome enablement of gar. Resources and genetic tools to be developed include: 1.) improvements in gar spawning techniques and infrastructure to perform molecular genetic experiments in gar; 2.) advances in CRISPR genome editing, targeted gene silencing, and transgenic technologies for the gar model; and 3.) dissemination of project results through a project-specific online database, two Gar Conferences, and two Workshops on Functional Vertebrate Evo-Devo Genomics. The project thereby seeks to understand the role of a select set of gar genes and regulatory elements as proof-of-principle to unlock the full potential of gars as model organisms for broad research questions in comparative vertebrate biology.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.