本文采用的英格恩产品: Entranster-H4000

Differential characteristics of mammalian and tick-derived promoters to trigger protein expression in transfected tick cell lines

Junming Shi  ¹ , Min Zhou  ¹ , Shuang Tang  ¹ , Qiaoli Wu  ¹ , Fei Deng  ¹ , Lesley Bell-Sakyi  ² , Shu Shen  ³ Affiliations

• PMID: 35114561
• DOI: 10.1016/j.ttbdis.2022.101906

Free article

Abstract

The transfection of plasmids into cell lines for the transient expression of exogenous proteins is a fundamental method for characterizing their functions, cellular localization and interactions. Currently, only a few reports on tick transfection systems and expression plasmids specifically constructed for tick cell lines have been published. In this study, the transcriptome of the tick cell line IDE8 was analyzed to screen for highly-expressed genes. The upstream sequences of these genes were selected as possible tick-derived promoters, and their promoter activity was evaluated using a luciferase assay. Four IDE8-derived sequences with promoter activity were identified, and the promoter activities of three common mammalian promoters, CMV, PGK and CAG, were studied and compared in the IDE8 and IRE/CTVM19 tick cell lines. In the two tick cell lines, the efficiency of the CAG promoter was considerably higher than that of CMV, PGK and the four newly-identified tick promoters. Additionally, time course experiments revealed that the protein expression driven by mammalian promoters reached peak levels on day 3, while the protein expression driven by our constructed tick-derived promoters reached peak levels on day 2 in tick cells. By comparing the transfection efficiency of three transfection reagents with different mechanisms in tick cell lines, we identified Effectene (with Enhancer, Qiagen) as the most effective reagent for tick cells. The findings of this study suggested that there are differences between tick and mammalian cell lines in their response to the transfection system. These findings will contribute to future studies on topics including tick protein function, tick genetic modification and tick-host-pathogen interactions.

Keywords: Luciferase assay; Tick cell lines; Tick-derived promoters; Transfection.