Direct generation of complex genetically-modified mouse models via embryonic stem cells
通过胚胎干细胞直接生成复杂的转基因小鼠模型
基本信息
- 批准号:10354630
- 负责人:
- 金额:$ 25.43万
- 依托单位:
- 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:2022-03-15 至 2024-02-28
- 项目状态:已结题
- 来源:
- 关键词:AllelesAnimal ModelAnimalsBiological ModelsBiologyBreedingBypassCRISPR/Cas technologyCattleCell Culture SystemCell Culture TechniquesChimera organismComplementComplexConsumptionDataDevelopmentDifferentiation InhibitorES Cell LineEmbryoEpigenetic ProcessExploratory/Developmental Grant for Diagnostic Cancer ImagingFemaleFosteringGenerationsGenesGeneticGenetically Modified AnimalsGenomeGenomic InstabilityGoalsInstitutesKnock-inLipidsMAPK3 geneMEK inhibitionMEKsMaintenanceMediatingMedical ResearchModificationMouse StrainsMusMutationPathway interactionsProcessProductionPublic HealthResearchResearch PersonnelRewardsSerumSerum-Free Culture MediaSignal PathwaySiteSpeedStem cell pluripotencySystemTechnologyTestingTimeUnited States National Institutes of HealthY ChromosomeY chromosome deletionbaseconditional knockoutcostembryonic stem cellflexibilitygenetic manipulationhigh rewardhigh riskhuman diseaseimprovedinhibitorinterestmalemouse modelmutantnovelpluripotencypreservationsexstemstem cellstooltransmission process
项目摘要
PROJECT SUMMARY: Genetically modified (GM) animals are essential tools for the
study of both fundamental biology and human diseases. The production of GM animals
relies on two critical technologies: 1) stable genetic modifications and 2) germline
transmission of the mutations into a model system. A typical approach for creation of
complex GM mice involves the generation of tetra-parental chimeras from normal
embryos and GM embryonic stem (ES) cells, followed by multiple rounds of breeding to
obtain both male and female mice for germline propagation. Two limitations dominate
this approach. First, maintenance of pluripotency limits the complexity of genetic
manipulations. Second, this process is time-consuming, laborious, and costly,
particularly if the final objective requires many independent germline manipulations in
the same animal. We propose a feasible strategy to accelerate the production of
complex GM mouse models. Employing the technology of sex-reversion via
CRISPR/Cas9-meditated Y chromosome deletion in male ES cells and our novel ES
cell culture system, we can directly generate isogenic male and female mice from the
same targeted ES cells through tetraploid complementation (4n). This strategy would
bypass at least two mouse breeding generations: the chimera development step and
the complex breeding process. We will target male (XY) ES cell lines for intended
genetic alterations and follow the deletion of Y chromosome to generate monosomic XO
female ES cells. Using this strategy, compound homozygous GM mouse strains could
be established at unparalleled speed and costs. In this R21 application we propose the
following two Aims: Aim 1: Optimize AX-based ES cell culture system for the production
of GM mouse models. Aim 2. Direct generation of isogenic male and female complex
GM mice using novel ES cell culture medium. If successful, our approach would have a
great impact on GM mouse model construction in terms of versatility, speed, and cost.
This ambitious endeavor to develop a breakthrough technology for creation of complex
GM mouse models would also possibly foster novel research opportunities.
项目摘要:转基因 (GM) 动物是实现人类健康的重要工具
研究基础生物学和人类疾病。转基因动物的生产
依赖两项关键技术:1) 稳定的基因修饰和 2) 种系
将突变传输到模型系统中。创建的典型方法
复杂的转基因小鼠涉及从正常小鼠产生四亲嵌合体
胚胎和转基因胚胎干(ES)细胞,然后经过多轮育种
获得雄性和雌性小鼠进行种系繁殖。主要有两个限制
这种方法。首先,多能性的维持限制了遗传的复杂性
操纵。其次,这个过程费时、费力、成本高,
特别是如果最终目标需要许多独立的种系操作
同一种动物。我们提出了一个可行的策略来加速生产
复杂的转基因小鼠模型。采用性别回复技术
CRISPR/Cas9介导的雄性ES细胞中Y染色体缺失和我们的新型ES
细胞培养系统,我们可以直接从细胞培养中产生同基因雄性和雌性小鼠
通过四倍体互补 (4n) 获得相同的目标 ES 细胞。这一策略将
绕过至少两代小鼠繁殖:嵌合体发育步骤和
复杂的养殖过程。我们将针对雄性 (XY) ES 细胞系进行预期
遗传改变并随着 Y 染色体的缺失产生单体 XO
雌性 ES 细胞。利用这种策略,复合纯合转基因小鼠品系可以
以无与伦比的速度和成本建立。在此 R21 应用中,我们建议
以下两个目标: 目标 1:优化基于 AX 的 ES 细胞培养系统用于生产
转基因小鼠模型。目标2.直接产生同基因雄性和雌性复合体
使用新型 ES 细胞培养基的转基因小鼠。如果成功的话,我们的方法将具有
在通用性、速度和成本方面对转基因小鼠模型构建产生了巨大影响。
这项雄心勃勃的努力旨在开发突破性技术来创造复杂的
转基因小鼠模型也可能带来新的研究机会。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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{{ truncateString('Duancheng Wen', 18)}}的其他基金
Direct generation of complex genetically-modified mouse models via embryonic stem cells
通过胚胎干细胞直接生成复杂的转基因小鼠模型
- 批准号:
10589018 - 财政年份:2022
- 资助金额:
$ 25.43万 - 项目类别:
H3.3-mediated epigenetic regulation of developmental bivalent genes for reprogramming and differentiation
H3.3介导的发育二价基因的表观遗传调控,用于重编程和分化
- 批准号:
10408113 - 财政年份:2018
- 资助金额:
$ 25.43万 - 项目类别:
H3.3-mediated epigenetic regulation of developmental bivalent genes for reprogramming and differentiation
H3.3介导的发育二价基因的表观遗传调控,用于重编程和分化
- 批准号:
10174958 - 财政年份:2018
- 资助金额:
$ 25.43万 - 项目类别:
H3.3-mediated epigenetic regulation of developmental bivalent genes for reprogramming and differentiation
H3.3介导的发育二价基因的表观遗传调控,用于重编程和分化
- 批准号:
9750724 - 财政年份:2018
- 资助金额:
$ 25.43万 - 项目类别:
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