GHS-R基因5'侧翼区萤火虫荧光素酶报告基因增强子载体的构建和鉴定

Construction and identification of the firefly luciferase reporter gene enhancer vectors for 5' flanking region of GHS-R gene

目的构建生长激素释放剂受体(GHS-R)基因5'侧翼区萤火虫荧光素酶报告基因增强子载体。方法以原代培养的人生长激素(GH)腺瘤细胞基因组DNA为模板,PCR扩增GHS-R基因5'侧翼转录调控区不同长度片段(A～F)克隆,再定向克隆到pGL3-Enhancer载体中。结果通过酶切鉴定和基因测序,证明成功地构建了pGL3-Enhancer A～F克隆重组体。结论本载体的成功构建为体外研究GHS-R基因的转录调节提供了新的手段,为下一步在人垂体GH腺瘤中GHS-R基因启动子的干预研究奠定了基础。

Objective To construct a series of firefly luciferase reporter gene enhancer vectors for 5 flanking region of GHS-R gene.Methods An A to F series of DNA fragments of 5 flanking transcription regulation region of GHS-R gene were amplified by PCR from the genomic DNA of primary cultured human pituitary somatotrophinomas cells.The PCR products were then cloned into the pGL3-Enhancer vector.Results Restriction enzyme digestion and nucleotide sequencing confirmed that the recombinant plasmids pGL3-Enhancer A-F...