Constructing the dual luciferase reporter vector containing human DRD1 promoter region

doi:10.3969/j.issn.2095-4344.2016.40.020

Abstract

Abstract:

BACKGROUND: The polymorphisms of dopamine receptor in promoter region will affect the expression of the receptor, thereby affecting the dopaminergic neurotransmitter, finally lead to related diseases.

OBJECTIVE: To construct the dual luciferase reporter vector containing human DRD1 promoter region and determine its activity, which could provide the basic tool for studying the transcriptional regulation of DRD1 gene.

METHODS: DRD1 promoter sequence was amplified by PCR using the human blood genomic DNA and cloned into pGM-T vector. After sequencing, the correctly constructed vectors were ligated to the firefly luciferase reporter plasmid pGL3-Basic. The cloned pGL3-Basic vectors were transfected into HEK293 using cationic liposome method. In the meanwhile, PGL3-Basic vector with no promoter was co-transfected with pGL3-TK plasmid as negative control group. The relative fluorescence intensity was measured by chemiluminescence.

RESULTS AND CONCLUSION: (1) Recombinant luciferase reporter gene vectors were confirmed by restriction analysis and sequencing. (2) Compared with the negative control group, the HEK293 cells transfected by recombinant vectors presented transcriptional activity. (3) In conclusion, luciferase reporter gene vectors containing DRD1 promoter region are successfully constructed and can provide the basic tool for further study on the transcriptional regulation of DRD1.

中国组织工程研究杂志出版内容重点：肾移植；肝移植；移植；心脏移植；组织移植；皮肤移植；皮瓣移植；血管移植；器官移植；组织工程

Key words: Receptors, Dopamine, Genes, Reporter, Transcription Initiation Site, Tissue Engineering

CLC Number:

R318

Wang Chun-hong, Li Zhe, Wang Ming-han, Deng Li-li, Wang Huan. Constructing the dual luciferase reporter vector containing human DRD1 promoter region[J]. Chinese Journal of Tissue Engineering Research, 2016, 20(40): 6060-6066.

Figures/Tables 2

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