Establishment of Somatic Cell Nuclear Transfer as a Universal Platform for Cloning Marmosets

体细胞核移植作为克隆狨猴通用平台的建立

基本信息

批准号：
10682479
负责人：
Jung Eun Park
金额：
$ 19.88万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-15 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10682479
关键词：
Address Anatomy Aneuploidy Animals Antimitotic Agents Biomedical Research Birth Brain CADASIL CRISPR/Cas technology Callithrix Callithrix jacchus jacchus Cell Nucleus Cell fusion Characteristics Chemicals Chromosomes Chronic Cloning Cognitive Combined Modality Therapy Communities Complex Crossbreeding Cytoplasm Cytosine DNA DNA Methylation DNA methyltransferase inhibition DNMT3B gene DNMT3a Deacetylase Development Domestic Animals Early Onset Alzheimer Disease Embryo Embryo Cloning Embryo Transfer Embryonic Development Epigenetic Process Female Funding Gene Expression Gene Expression Profile Gene Transfer Techniques Generations Genes Genetic Genetic Engineering Genetic study Goals Histone Acetylation Histones Human In Vitro Individual Infant Measures Mechanics Messenger RNA Metabolic Metaphase Methods Methyltransferase Microvascular Dysfunction Modeling Mosaicism Mus Mutation NOTCH3 gene Neurosciences Research Nuclear Oocytes Ooplasm Phenotype Phylogenetic Analysis Physiological Pregnancy Procedures Process Protocols documentation RNA Research Research Support Social Behavior Somatic Cell Stains System Techniques Technology Term Birth Testing Totipotent Transgenes Transgenic Organisms Translational Research Ultraviolet Rays United States National Institutes of Health Variant Work assistive reproductive technique blastocyst calcium indicator efficacy evaluation epigenome experience genome editing histone demethylase human disease human model immune function improved inhibitor interest mosaic nervous system disorder neuropsychiatric disorder nonhuman primate novel nuclease presenilin-1 preservation prevent reproductive somatic cell nuclear transfer tool translational neuroscience

项目摘要

PROJECT SUMMARY/ABSTRACT The common marmoset (Callithrix jacchus) is a New World non-human primate (NHP) with several practical advantages in biomedical research. Due to their phylogenetic proximity, marmosets are a genetically diverse NHP species with similar physiological, metabolic, and immunological functions as humans. Marmosets retain the typical anatomical and functional organization of the human brain. Marmosets have complex cognitive and social behavior. The above characteristics place marmosets as an ideal NHP model to bridge the gap between mice and humans for both basic and translational neuroscience. Marmosets reach sexual maturity at circa 18 months and give birth to multiple infants twice a year. Their short gestation period and compatibility with gene editing techniques make marmosets ideally poised to become the NHP model of choice in studying the genetic causes of neurological and neuropsychiatric disorders and understanding brain function. We have successfully generated transgenic marmosets expressing genetically encoded calcium indicators and genetically engineered marmosets with NOTCH3 mutations that cause the small vessel disease CADASIL. We have also made marmosets harboring PSEN1 mutations that cause early-onset Alzheimer’s disease. Our gene-edited marmosets will enable us to investigate the genetic causes of chronic neurological disorders. However, two main issues hinder the broader availability of genetically engineered marmosets. First, lentiviral-based approaches suffer from an uncontrollable integration of transgene with variation in copy number, and nuclease-based gene editing produces embryos with mosaic editing, leading to unpredictable gene expression patterns and variable phenotype. Second, and most importantly, there are no efficient ways to propagate an individual showing the interest phenotype. This proposal addresses these shortcomings by developing and optimizing somatic cell nuclear transfer (SCNT) as a universal platform for cloning marmosets. We want to build on our vast experience generating genetically modified marmosets to (1) develop and optimize the enucleation procedure in marmoset somatic cell nuclear transfer and (2) develop and optimize approaches enabling technologies for efficient epigenetic reprogramming during somatic cell nuclear transfer in the marmoset. These results will establish optimized marmoset SCNT protocols and unravel a novel enabling universal platform to generate a sizeable number of cloned marmosets expressing any phenotype of interest. This platform will significantly facilitate the propagation and sharing of marmoset models for neuroscience and translational research supported by multiple NIH ICs.

项目摘要/摘要普通的摩尔果（Callithrix jacchus）是一个新世界非人类灵长类动物（NHP），有几个实用性生物医学研究的优势。由于其系统发育近端具有类似生理，代谢和免疫学功能的NHP物种与人类相似。马尔莫斯特保留人脑的典型解剖和功能组织。果果具有复杂的认知和社会行为。以上特征将摩尔马人作为理想的NHP模型，以弥合差距小鼠和人类的基本和转化神经科学。大约18岁的马尔莫斯人达到性成熟几个月，每年两次生下多名婴儿。他们的短期妊娠期和与基因的兼容性编辑技术使Marmosets理想中毒成为研究通用的NHP模型神经和神经精神疾病的原因并了解大脑功能。我们已经成功产生的转基因摩尔果会表达一般编码的钙指标和基因工程带有NOTCH3突变的摩尔果会导致小血管疾病CADASIL。我们也做了带有PSEN1突变的摩尔果会引起早期发作的阿尔茨海默氏病。我们的基因编辑 Marmosets将使我们能够研究慢性神经系统疾病的遗传原因。但是，两个主要问题阻碍了一般设计的果酱的广播公司的可用性。首先，基于慢病毒的方法遭受转化不可控制的整合与拷贝数的变化和基于核酸酶的基因的变化编辑产生带有镶嵌编辑的胚胎，导致不可预测的基因表达模式和可变表型。其次，最重要的是，没有有效的方法来传播一个个人兴趣表型。该建议通过开发和优化体细胞来解决这些缺点核转移（SCNT）是克隆果棒的通用平台。我们想建立我们的丰富经验在（1）中生成一定经过修改的Marmosets，并在Marmoset中开发和优化枚举过程体细胞核转移和（2）开发和优化实现技术以提高技术的方法在马尔莫斯群体中的体细胞核转移过程中表观遗传重编程。这些结果将确定优化的Marmoset SCNT协议，并解散了一种新颖的使通用平台能够生成大量的平台表达任何感兴趣表型的克隆果果的数量。该平台将大大促进由多重支持 NIH IC。