Acta Entomologica Sinica ›› 2019, Vol. 62 ›› Issue (4): 391-397.doi: 10.16380/j.kcxb.2019.04.001

• RESEARCH PAPERS •     Next Articles

Construction of a simple basal promoter for the research of transcription regulatory elements in the silkworm (Bombyx mori) at the cell level

 GU Jian-Jian1,2, LIU Hong-Ling1,2, LI Kai-Rong2,3, MENG Zi-Wang2,3, WANG Hui-Juan2,3, SHEN Guan-Wang1,2, ZHANG Yu-Jing1,2, RUAN Yang1,2, LIN Ying1,2,3,*, XIA Qing-You1,2,3   

  1. (1. State Key Laboratory of Silkworm Genome Biology,SouthwestUniversity,Chongqing400716,China; 2.ChongqingKey Laboratory of Sericultural Science,Chongqing400716,China;  3.CollegeofBiotechnology,SouthwestUniversity,Chongqing400716,China)
  • Online:2019-04-20 Published:2019-04-08

Abstract: Aim This study aims to construct a simple basic promoter, which can be stably expressed in the silkworm (Bombyx mori) cells, so as to reflect the influence of single transcriptional regulatory element on gene promoter activity more accurately and lay a foundation for studying the transcriptional regulation of the silkworm and even other insects. Methods Based on the BmVgP78Mpromoter previously reported in our research group, which can be stably expressed in silkworm cells and hardly contains upstream transcriptional regulatory elements, interval sequences of a certain length and BrC-Z2 transcription factor binding motif (BrC-Z2 element, BrC-Z2E) that can respond to 20-hydroxyecdysone (20E) and enhance promoter activity were added to the upstream of BmVgP78Mby PCR technique. The cell transfection vectors were constructed by gene cloning technique, and the changes of promoter activity were detected by cell transfection technique and dual luciferase reporter gene system. Results A simple basal promoter named VgP78ML was constructed successfully with a 28 bp interval sequence added to the upstream of the BmVgP78Mpromoter, and proved to be a simple basic promoter to study target transcriptional regulatory elements. The verification test results showed that this simple basal promoter not only could be stably expressed in silkworm cells, but also was not affected by 20E and transcription factor BrC-Z2. When BrC-Z2E was linked to the upstream of this promoter, it could significantly respond to 20E and BrC-Z2 transcription factor, thereby regulating the expression of reporter gene. Conclusion VgP78ML can be used as a simple basic promoter to study the transcriptional regulation of genes in silkworm at the cell level. At the same time, its construction method also provides a reference for constructing simple basic promoter to study the transcriptional regulation in other species.

Key words: Bombyx mori; cell, simple basal promoter, dual luciferase reporter gene system, transcriptional regulation, transcription element