Construction of recombinant plasmid containing human SOX18 gene and effect of the protein on proliferation of vascular endothelial cells
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摘要:
目的：构建含人SOX（SRY related high mobility group box）18的重组表达质粒，并研究其对血管内皮细胞增殖能力的影响。方法：通过RT-PCR技术从血管内皮细胞中扩增SOX18编码基因序列，将其插入真核表达载体pSecTag-2B构建重组表达质粒pST-SOX18，分别转染人胚肾上皮细胞HEK293T和血管内皮细胞，通过免疫印迹技术检测SOX18的蛋白表达情况；进一步通过CCK-8实验检测SOX18对血管内皮细胞增殖能力的影响。结果：成功构建重组质粒pST-SOX18，重组蛋白能够在HEK293T和血管内皮细胞中表达；SOX18的过表达促使血管内皮细胞合成血管内皮生长因子（VEGF），促进血管内皮细胞增殖。结论：SOX18重组蛋白能够促进血管内皮细胞增殖。
Abstract:
Objective：To construct the recombinant plasmid containing SRY related high mobility group box 18（SOX18）gene and investigate the effect of the recombinant protein on angiogenesis of vascular endothelial cells. Methods：The whole sequence of SOX18 gene that amplified from human umbilical vein endothelial cells（HUVEC）by reverse transcription-polymerase chain reaction（RT-PCR）were inserted into the expression plasmid pSecTag-2B to construct the recombinant vector，and termed as pST-SOX18. pST-SOX18 was then transfected into the HEK293T and vascular endothelial cells，respectively. Subsequently，Western blotting was carried out to determine the expression of the recombinant protein. CCK-8 assay was used to assess the effect of pST-SOX18 on proliferation of vascular endothelial cells. Results：The recombinant plasmid pST-SOX18 was constructed successfully. pST-SOX18 was successfully transfected into HEK293T and vascular endothelial cells，target protein was determined by immunoblotting. pST-SOX18 transfection significantly promoted vascular endothelial cell proliferation in vitro. Conclusion：The recombinant plasmid expressing functional SOX18 was constructed successfully. SOX18 have a potent potential in promoting the proliferation of vascular endothelial cells.
