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CALCITONIN RECEPTORS IN DEVELOPMENT

CALCITONIN RECEPTORS IN DEVELOPMENT
降钙素受体的发育
批准号:
6100314
负责人:
STEVEN GOLDRING
金额:
$0.0万
依托单位国家:
美国
项目类别:
财政年份:
1997
资助国家:
美国
项目状态:
已结题
起止时间:
1997-01-01 至 1998-12-31

项目摘要

项目成果

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中文摘要
翻译
降钙素(CT)最初被确定为一种肽激素释放 甲状腺滤泡旁细胞对高钙反应 刺激。 在哺乳动物中,发现CT分泌产生快速的 通过抑制骨吸收降低细胞外钙水平 和刺激肾钙排泄,高钙介导的作用, 亲和受体位于破骨细胞和某些肾小管 细胞 随后,研究表明CTR分布广泛, 提示CT可能具有更复杂和多样的功能。 CT可以 在脊椎动物胚胎的早期分化中也很重要。 本建议的目的是完成确定 结构的小鼠CTR基因,引入靶向突变, 基因在胚胎干细胞中同源重组, 表达这些突变的转基因小鼠, CT和CTR基因在胚胎发育中的作用。 斑马鱼系统 也将用于了解CTR在早期胚胎发育中的作用。 发展 在具体目标1中,确定化合物的结构 将完成鼠CTR基因,并将5 '-调控序列 鉴定 在本实验室测序的小鼠脑CTR cDNA, 将用于鉴定CTR基因中的外显子(见项目4)。 在 特异性目的2,从鼠CTR cDNA制备的限制性片段 或者将基因组CTR克隆亚克隆到载体中以产生CTR- 特异性cRNA探针 这些将用于CTR基因的定位 转录在不同阶段的小鼠发展与技术 原位杂交技术。 CTR表达的模式通过以下测定: 原位杂交将与通过以下方法获得的结果相关: 放射性碘标记CT放射自显影和CTR免疫组化 抗体的 在特定目标3中,胚胎中的同源重组 茎(ES)将用于CTR基因的靶向破坏; ES 细胞将用于生产转基因小鼠。 这将允许直接 在鼠中破坏CTR基因的后果的测定 胚胎发育。 在特定目标4中,lacZ基因将被放置在 就在CTR基因翻译起始位点的下游, 通过ES细胞同源重组产生突变小鼠。 这种方法将允许在体内确定空间和 CTR基因的时间表达和表达CTR基因的细胞谱系 基因在发育过程中 在特定目标5中,斑马鱼CTR cDNA将 克隆这些序列或来自本发明的序列的载体, CTR基因将用于检查异位妊娠的后果。 CT或CTR在早期胚胎中的产生或过度表达 发展
英文摘要
Calcitonin (CT) was originally identified as a peptide hormone released by parafollicular cells of the thyroid gland in response to hypercalcemic stimuli. In mammals, it was found that CT secretion produces a rapid lowering of extracellular calcium levels by inhibition of bone resorption and stimulation of renal calcium excretion, effects mediated by high affinity receptors located on osteoclasts and certain renal tubular cells. Subsequently, it was shown that CTRs are widely distributed, suggesting that CT may have more complex and diverse functions. CT may also be important in the early differentiation of the vertebrate embryo. The aims of this proposal are to complete the determination of the structure of the murine CTR gene, introduce targeted mutations into the gene by homologous recombination in embryonic stem cells and create transgenic mice that express these mutations in order to define the role of CT and the CTR gene in embryonic development. The zebrafish system will also be used to understand the role of the CTR in early embryonic development. In Specific Aim 1, determination of the structure of the murine CTR gene will be completed and the 5'-regulatory sequences identified. The murine brain CTR cDNA, sequenced in this laboratory, will be used to identify exons in the CTR gene (see Project 4). In Specific Aim 2, restriction fragments prepared from the murine CTR cDNA or the genomic CTR clones will be subcloned into vectors to produce CTR- specific cRNA probes. These will be utilized to localize CTR gene transcripts during various stages of murine development with techniques of in situ hybridization. The pattern of CTR expression determined by in situ hybridization will be correlated with that obtained by autoradiography with radioiodinated CT and immunohistochemistry with CTR antibodies. In Specific Aim 3, homologous recombination in embryonic stem (ES) will be used for targeted disruption of the CTR gene; the ES cells will be used to produce transgenic mice. This will permit direct determination of the consequences of disruption of the CTR gene in murine embryonic development. In Specific Aim 4, the lacZ gene will be placed just downstream from the translation initiation site in the CTR gene by means of homologous recombination of ES cells to produce mutant mice. This approach will permit the determination in vivo of the spatial and temporal expression of the CTR gene and the lineage of cells that express the gene during development. In Specific Aim 5, zebrafish CTR cDNA will be cloned and vectors incorporating these sequences or sequences from the CTR gene will be utilized to examine the consequences of ectopic production or overexpression of CT or the CTR in early embryonic development.
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  • 财政年份:
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  • 负责人:
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  • 依托单位:
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  • 项目类别:
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国内基金
海外基金
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  • 项目类别:
    面上项目
  • 资助金额:
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  • 批准年份:
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  • 依托单位: