Establishment of a suitable control reporter plasmid of a dual luciferase reporter gene system for hormone research in silkworm cell lines.
- Author:
Hongling LIU
1
;
Ying LIN
1
;
Guanwang SHEN
1
;
Jianjian GU
1
;
Haiyan ZHANG
1
;
Jinxin WU
1
;
Yinying XU
1
;
Wei LONG
1
;
Qingyou XIA
1
Author Information
- Publication Type:Journal Article
- Keywords: control reporter plasmid; dual luciferase reporter gene system; hormone; method; silkworm
- From: Chinese Journal of Biotechnology 2018;34(10):1631-1641
- CountryChina
- Language:Chinese
- Abstract: The dual luciferase reporter gene system provides sensitive readout, while it relies on a constitutively-expressed control gene for readout normalization. However, most standard control reporter genes are not constitutively expressed under all conditions. Here, we report an effective method to construct a control reporter plasmid for the dual luciferase reporter gene system that would be suitable for hormone research in silkworm cell lines. First, we modified BmVgP78M, a stably-expressed constitutive promoter in silkworm cells by mutating its hormone-related element. Then, we constructed the pRL-VgP78M control reporter plasmid by replacing the SV40 promoter and chimeric intron sequences in pRL-SV40 with the BmVgP78M sequence. Finally, we confirmed that the pRL-VgP78M control reporter plasmid could be stably expressed in silkworm cell lines via cell transfection experiments, and it was unresponsive to the induction of ecdysone, juvenile hormone, or their transcription factors. We thus obtained a control reporter plasmid pRL-VgP78M that could be expressed stably and moderately in silkworm cells. It can be readily used as the control reporter plasmid of the dual luciferase reporter gene system for hormone research in silkworm cell lines. It will also provide a reference for construction of control reporter plasmids of dual luciferase reporter gene systems that are adaptable to cell lines isolated from other species.
