Related Experiment Video
Updated: Jun 17, 2026
Generation of Transgenic Rats using a Lentiviral Vector Approach
Published on: May 17, 2020
[Construction and identification of eukaryotic expression vector of rat GPR17 gene]
Ka-Na Lin1, San-Hua Fang, Bei-Lei Cai
1Department of Pharmacology, College of Medicine, Zhejiang University, Hangzhou 310058, China.
Objective:
To construct the eukaryotic expression vector of rat GPR17 (rGPR17) cDNA,and to identify its function in HEK293 cells.
Methods:
Total RNA was extracted from rat brain tissue; full-length GPR17 cDNA was prepared by RT-PCR, and cloned into pcDNA3.1(+) plasmid. The recombinant plasmid was converted into E.coli DH5alpha and confirmed by PCR, double enzyme digestion analysis and DNA sequencing. The recombinant plasmid pcDNA3.1(+)-rGPR17 was transiently transfected into HEK293 cells using Lipofectamin 2000. Expression of rGPR17 gene was confirmed by RT-PCR and immunofluorescence staining. The exogenous LTD(4) enhanced intracellular calcium was measured using Fluo-4.
Result:
RT-PCR, double enzyme digestion analysis and sequencing showed that the rGPR17 gene was cloned into recombinant vector, and the recombinant rGPR17 was expressed after transfection in HKE293 cells. LTD(4) increased intracellular calcium release in the transfected HEK293 cells.
Conclusion:
The eukaryotic expression vector of rGPR17 cDNA has been constructed; it is functionally expressed in HEK293 cells. This work provides a basis for further research of the GPR17 receptor and its antagonists.
More Related Videos
04:18A Seamless Cloning Approach for Porcine Reproductive and Respiratory Syndrome Virus Expression Vector Construction
Published on: May 17, 2024
12:48Construction of Cell-based Neurotransmitter Fluorescent Engineered Reporters (CNiFERs) for Optical Detection of Neurotransmitters In Vivo
Published on: May 12, 2016