转录因子转录活性检测

服务介绍

转录因子根据其功能的不同可以分为转录激活因子和转录抑制因子，转录激活能够促进基因转录，而转录抑制因子能够抑制基因转录。转录因子可以通过多种方式影响基因转录，其中转录激活因子能够与RNA聚合酶和其它调节因子形成复合物，通过与DNA结合并改变其染色质状态，从而使RNA聚合酶能够更容易地接近基因启动子区域，进而促进基因的转录；还有一些转录因子则通过招募转录抑制因子，从而抑制基因的转录。

酵母检测系统：

单独的BD可以与GAL4上游活化序列UAS结合，但不能引起转录，但是如果把一个具有转录激活活性的转录因子构建到BD载体上，其表达产生的诱饵蛋白单独与UAS结合后能够引起下游报告基因的转录和表达，根据下游报告基因的转录和表达情况我们就可以判断转录因子是否具有转录激活活性。

双荧光素酶报告基因检测系统：

将待测转录因子与GAL4结合域构建在同一载体上，与含有GAL-TATA转录调控原件并带有荧光虫荧光素酶(Firefly luciferase)的表达载体共转，使这段序列调控luciferase的转录表达。然后将报告基因质粒转染细胞，培养后裂解细胞，并加入底物荧光素，荧光素酶可催化荧光素发出荧光。通过检测荧光值的高低可以判断转录因子是否具有转录激活或者转录抑制作用。

服务优势

▶实验结果可靠、图片美观；

▶除了转录激活检测实验外，还可以提供酵母互作以及文库筛选等业务，可以进一步丰富实验内容。

服务流程

 酵母检测系统

双荧光素酶报告基因检测系统

文案展示

(1) 酵母检测系统

Natural variation of OsWRKY23 drives difference in nitrate use efficiency between indica and japonica rice

期刊：nature communications

OsWRKY23 showed transactivation activity in yeast. Full-length, N-terminal deletion, and C-terminal deletion cDNAs of OsWRKY23 were cloned into pGBKT7 and fused with the GAL4 DNA-binding domain. Numbers indicate the positions of the amino acid of OsWRKY23.  

(2) 双荧光素酶报告基因检测系统

Plant NAC-type transcription factor proteins contain a NARD domain for repression of transcriptional activation

期刊：Planta 

EVect of NARD on VP16 activity in protoplast assay. a Schematic representation of constructs used in Arabidopsis transient assay. The reporter construct GAL4-LUC contained Wve copies of the GAL4 responsive element, minimal TATA box region (starting at position ¡46) of 35S promoter, the WreXy gene for luciferase (LUC; shown as a closed box), and a nopaline synthase (Nos) terminator. Each of the eVectors contained a GAL4 DNA-binding domain (GAL4DBD). GALDBD was also used as a negative control. The GAL4DBD was fused with VP16 (VP16), or with two copies of VP16 (VP16 VP16) and used as positive controls. NARD was fused to the N-terminal or C-terminal of VP16 to test its eVects. All the constructs were driven by 35S promoter with translation enhancer sequence, and ended by Nos terminal. b NARD inhibits the VP16 activity. Relative luciferase activity was measured after PEG co-transfection of protoplasts with GAL4-LUC reporter and GAL4 fusion eVector plasmids. Luciferase activity was examined when the transformed protoplasts were cultured for 6 h.

适用领域

▶转录因子对下游基因的作用。

▶验证转录因子活性。