腭裂相关基因甲状腺转录因子-2转基因小鼠的生物学特征

发布时间：2018-05-13 05:09

本文选题：甲状腺转录因子- + 持续表达　；参考：《华西口腔医学杂志》2014年04期

【摘要】：目的建立腭裂相关基因甲状腺转录因子-2(TTF-2)转基因小鼠模型,利用转基因动物模型研究TTF-2基因的活动规律和表达模式发生变化时对腭突发育过程产生的影响。方法采用聚合酶链反应法扩增C57BL/6J小鼠基因组TTF-2基因,将其定向插入pBROAD3-mcs载体,构建重组表达质粒pBROAD3-TTF-2,利用显微注射技术把线性化表达载体注射到受精卵的雄原核中,建立TTF-2转基因小鼠。利用特异引物聚合酶链反应和Southern blot方法鉴定转基因小鼠的基因型,采用免疫组织化学法检测TTF-2基因在转基因小鼠腭突组织中的表达。结果共注射原核期受精卵982枚,发育为2-细胞胚胎的580枚,均移植入48个假孕受体昆明白小鼠中,得到胎鼠68只。提取基因组DNA,发现有13只转基因阳性的胎鼠。免疫组织化学法检测显示TTF-2在转基因小鼠腭突中持续表达。结论通过显微注射法使外源基因pBROAD3-TTF-2整合入小鼠基因组中,成功建立了腭突持续表达TTF-2的转基因小鼠模型,其表现型为腭裂。
[Abstract]:Objective to establish a transgenic mouse model of cleft palate related gene thyroid transcription factor-2 TTF-2, and to study the effect of TTF-2 gene activity and expression pattern on the development of palatine process by transgenic animal model. Methods the genomic TTF-2 gene of C57BL/6J mice was amplified by polymerase chain reaction and inserted into the pBROAD3-mcs vector. The recombinant expression plasmid pBROAD3-TTF-2 was constructed. The linearized expression vector was injected into the male prokaryotic cells of fertilized eggs by microinjection technique. TTF-2 transgenic mice were established. The genotypes of transgenic mice were identified by specific primer polymerase chain reaction (PCR) and Southern blot method. The expression of TTF-2 gene in palatine process of transgenic mice was detected by immunohistochemistry. Results A total of 982 fertilized eggs were injected into Kunming white mice with 48 pseudopregnant receptors, and 68 fetal mice were obtained. Genomic DNA was extracted and 13 transgenic fetal mice were found. Immunohistochemical staining showed that TTF-2 was expressed continuously in the palatal process of transgenic mice. Conclusion the transgenic mouse model of persistent expression of TTF-2 in palatine process was successfully established by microinjection of exogenous gene pBROAD3-TTF-2 into the mouse genome, and the phenotype was cleft palate.
【作者单位】：广州市第一人民医院口腔科;口腔疾病研究国家重点实验室华西口腔医院(四川大学);口腔疾病研究国家重点实验室华西口腔医院(四川大学)唇腭裂外科;
【基金】：国家自然科学基金资助项目(30171020)
【分类号】：R782.2

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