| 叶忠张必利# 徐荣良赵仙先秦永文唐念中郑兴△.斑马鱼心肌特异性启动子的克隆及其功能鉴定*[J].,2012,12(27):5204-5208 |
| 斑马鱼心肌特异性启动子的克隆及其功能鉴定* |
| Cloning and Functional Identification of the Zebrafish Cardiac-specificPromoter* |
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| DOI： |
| 中文关键词: 斑马鱼 vmhc 基因 启动子 显微注射 |
| 英文关键词: Zebrafish Vmhc gene Promoter Microinjection |
| 基金项目:国家自然科学基金项目（81000038） |
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| 中文摘要: |
| 目的：探讨心室肌球蛋白重链（vmhc）基因启动子的心肌组织特异性。方法：利用PCR 技术从斑马鱼基因组中克隆了vmhc
编码区5' 上游大小为1952bp 的调控区域，应用酶切连接方法将vmhc 启动子插入pGEFP-N1 质粒，成功构建pEGFP-vmhc 重组
载体。再应用高保真DNA 聚合酶PCR 扩增包含vmhc 启动子序列，增强型绿色荧光蛋白（EGFP）基因序列及3'UTR 序列的基因
片段，经过纯化后通过显微注射将vmhc-EGFP 基因片段导入斑马鱼受精卵中。结果：注射后的斑马鱼心脏中出现绿色荧光，而其
他部位无荧光出现。结论：vmhc 启动子能够正确有效地驱动外源基因在斑马鱼心脏中特异表达，适合应用于心血管疾病的基因
功能研究，基因靶向治疗等。 |
| 英文摘要: |
| Objective: To study the cardiac-specific expression of ventricular myosin heavy chain （vmhc） gene promoter.
promoter．Methods: A 1952 bp promoter fragment of zebrafish vmhc gene 5'end deletion upstream region was amplified by PCR from
zebrafish．Vmhc promoter was inserted into pGEFP-N1plasmid by enzyme digestion to construct the recombinant vector of
pEGFP-vmhc. The gene fragment contain vmhc promoter sequences,enhanced green fluorescent protein gene （EGFP）sequences and
3'UTR sequences was amplified by PCR with high-fidelity DNA polymerase. The purified vmhc-EGFP gene fragment was injected into
fertilized eggs of zebrafish by Microinjection. Results: Green fluorescence could be observed in the zebrafish heart injected the
vmhc-EGFP gene fragment, but not in Non-myocardial tissue. Conclusion: Vmhc promoter is able to correctly and effectively drive the
specific expression of exogenous genes in the zebrafish heart, and it is suitable for the study of gene function and gene therapy in
cardiovascular disease. |
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