中国医学科学院学报
中國醫學科學院學報
중국의학과학원학보
ACTA ACADEMIAE MEDICINAE SINICAE
2009年
6期
692-695
,共4页
Setd7%神经分化%Ngn1
Setd7%神經分化%Ngn1
Setd7%신경분화%Ngn1
Setd7%neuron differentiation%Ngn1
目的 构建小鼠组蛋白赖氨酸甲基转移酶Setd7真核表达载体并初步探讨其对神经细胞分化的影响.方法 采用基因工程技术,克隆小鼠Setd7基因并将其插入真核表达载体pCMV-3tag-6中,之后转染HEK 293T细胞,Western blot验证其表达;利用实时荧光定量PCR技术检测在神经分化模型(全反式维甲酸诱导P19神经分化)中,Setd7对神经分化关键基因Ngn1的调控;利用双荧光素酶报告系统检测Setd7对Ngn1启动子活性的影响;构建小鼠Setd7 siRNA干扰质粒,利用实时荧光定量PCR技术检测其抑制效果以及对 Ngn1 mRNA表达的影响.结果 成功构建了小鼠Setd7基因的真核表达载体,可在HEK 293T细胞中有效表达;Setd7可促进Ngn1 mRNA表达;Setd7能够促进Ngn1启动子活性; 降低Setd7抑制Ngn1 mRNA 表达.结论 组蛋白赖氨酸甲基转移酶Setd7能够促进神经分化关键基因Ngn1的转录.
目的 構建小鼠組蛋白賴氨痠甲基轉移酶Setd7真覈錶達載體併初步探討其對神經細胞分化的影響.方法 採用基因工程技術,剋隆小鼠Setd7基因併將其插入真覈錶達載體pCMV-3tag-6中,之後轉染HEK 293T細胞,Western blot驗證其錶達;利用實時熒光定量PCR技術檢測在神經分化模型(全反式維甲痠誘導P19神經分化)中,Setd7對神經分化關鍵基因Ngn1的調控;利用雙熒光素酶報告繫統檢測Setd7對Ngn1啟動子活性的影響;構建小鼠Setd7 siRNA榦擾質粒,利用實時熒光定量PCR技術檢測其抑製效果以及對 Ngn1 mRNA錶達的影響.結果 成功構建瞭小鼠Setd7基因的真覈錶達載體,可在HEK 293T細胞中有效錶達;Setd7可促進Ngn1 mRNA錶達;Setd7能夠促進Ngn1啟動子活性; 降低Setd7抑製Ngn1 mRNA 錶達.結論 組蛋白賴氨痠甲基轉移酶Setd7能夠促進神經分化關鍵基因Ngn1的轉錄.
목적 구건소서조단백뢰안산갑기전이매Setd7진핵표체재체병초보탐토기대신경세포분화적영향.방법 채용기인공정기술,극륭소서Setd7기인병장기삽입진핵표체재체pCMV-3tag-6중,지후전염HEK 293T세포,Western blot험증기표체;이용실시형광정량PCR기술검측재신경분화모형(전반식유갑산유도P19신경분화)중,Setd7대신경분화관건기인Ngn1적조공;이용쌍형광소매보고계통검측Setd7대Ngn1계동자활성적영향;구건소서Setd7 siRNA간우질립,이용실시형광정량PCR기술검측기억제효과이급대 Ngn1 mRNA표체적영향.결과 성공구건료소서Setd7기인적진핵표체재체,가재HEK 293T세포중유효표체;Setd7가촉진Ngn1 mRNA표체;Setd7능구촉진Ngn1계동자활성; 강저Setd7억제Ngn1 mRNA 표체.결론 조단백뢰안산갑기전이매Setd7능구촉진신경분화관건기인Ngn1적전록.
Objective To construct the eukaryotic expression plasmid of mouse histone lysine methyltransferase Setd7 and detect its effect on neuron development. MethodsThe clone of mouse Setd7 was obtained and inserted into the eukaryotic expression vector pCMV-3tag-6-Flag. The plasmid was transfected into HEK 293T and identified by Western blot. Real-time PCR was used to detect the effect of Setd7 on the neuron differentiation marker gene Ngn1 mRNA expression. Dual luciferase reporter system was used to detect the effect of Setd7 on Ngn1 mRNA expression. Real-time PCR was used to detect the effect of Setd7 siRNA plasmid on Ngn1 mRNA expression. ResultsAn eukaryotic expression plasmid of Setd7 was successfully constructed. Setd7 induced Ngn1 mRNA expression and increased Ngn1 promoter activity. Also, the knockdown of Setd7 inhibited Ngn1 mRNA expression. ConclusionHistone lysine methyltransferase Setd7 can enhance neuron differentiation marker gene Ngn1 transcription.
