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目的:构建带myc标签的人EYA3基因真核表达载体,获得myc-EYA3融合表达蛋白,并对其功能进行初步检测。方法:采用PCR技术从乳腺文库中扩增人EYA3基因,并将其正确插入pXJ-40-myc载体;将重组质粒与空载体分别转染人乳腺癌细胞系ZR75-1后,Western印迹检测表达情况,并进行生长曲线实验。结果:双酶切和测序鉴定表明,myc-EYA3真核表达质粒构建成功,转染ZR75-1细胞后成功表达;生长曲线实验结果表明,EYA3可促进乳腺癌细胞的生长。结论:构建了带myc标签的人EYA3基因真核表达载体,myc-EYA3能在乳腺癌细胞ZR75-1中表达,且能促进该细胞的生长,本实验为进一步研究在乳腺癌中的功能奠定了基础。
OBJECTIVE: To construct eukaryotic expression vector of human EYA3 gene with myc tag and obtain myc-EYA3 fusion protein, and to detect its function. Methods: The human EYA3 gene was amplified from the mammary gland by polymerase chain reaction (PCR) and inserted into pXJ-40-myc vector. The recombinant plasmid and empty vector were transfected into human breast cancer cell line ZR75-1. Situation, and the growth curve experiment. Results: Double enzyme digestion and sequencing showed that myc-EYA3 eukaryotic expression plasmid was successfully constructed and transfected into ZR75-1 cells successfully. The growth curve results showed that EYA3 could promote the growth of breast cancer cells. CONCLUSION: The eukaryotic expression vector of human EYA3 gene with myc tag was constructed. Myc-EYA3 can be expressed in breast cancer cell ZR75-1 and promote the growth of the cell. This study is to further study the function of EYA3 gene in breast cancer The foundation.