CORE--TRANSGENIC MOUSE/GENE TARGETING

转基因小鼠/基因靶向核心将产生转基因小鼠， 原核注射携带特异性调节因子的基因构建体 在软骨和骨中表达的序列，与报告基因融合， 基因 转基因小鼠的创始人将被交付给个人 程序调查员进行后续分析。 四个项目中的三个 提出用一系列基因构建体产生转基因小鼠。 在 此外，这个核心将产生携带突变的ES细胞系， 软骨和骨骼发育基因的研究。 由该计划研究人员产生的基因靶向构建体将被 电穿孔到小鼠ES细胞中并在选择性条件下培养 以富集同源重组事件。 产生的ES菌落 将被挑选，扩大，并暂时冻结和DNA分裂 准备. DNA将被隔离并限制在适当的 内切酶 受限制的DNA随后将被交付给项目 研究人员随后进行了南方分析。 一旦确定，核心 将扩增目标克隆并在氮气中冷冻。 芯会 将携带靶向突变的ES克隆注射到胚泡中， 产生小鼠嵌合体。 由此产生的小鼠嵌合体将被递送到 以供后续分析。 中的每 这四个项目建议培育含有靶向突变的小鼠。 转基因小鼠/基因靶向核心代表了一个重要组成部分， 这个项目的计划。

The Transgenic mouse/gene targeting core will generate transgenic mice by pronuclear injection of gene constructs carrying specific regulatory sequences that are expressed in cartilage and bone, fused to a reporter gene. The transgenic mouse founders will be delivered to the individual Program investigators for subsequent analysis. Three of the four projects propose to generate transgenic mice with a series of gene constructs. In addition, this core will generate ES cell lines carrying mutations in the cartilage and bone development genes studied by the Program investigators. Gene targeting constructs generated by the Program investigators will be electroporated into mouse ES cells and cultured under selective conditions to enrich for homologous recombination events. The resulting ES colonies will be picked, expanded, and split for temporary freeze and DNA preparation. DNAs will be isolated and restricted with the appropriate enzymes. The restricted DNAs will then be delivered to the Project investigators for subsequent Southern analysis. Once identified, the core will expand targeted clones for freeze down in nitrogen. The core will inject ES clones carrying the targeted mutations into blastocysts to generate mouse chimeras. The resulting mouse chimeras will be delivered to the individual program investigators for subsequent analysis. Each of the four projects proposes to generate mice containing targeted mutations. The Transgenic mouse/gene targeting core represents an essential component of this Program Project.