Tobacco rattle virus-induced PHYTOENE DESATURASE (PDS) and Mg-chelatase H subunit (ChlH) gene silencing in Solanum pseudocapsicum L

Hua Xu; Leifeng Xu; Panpan Yang; Yuwei Cao; Yuchao Tang; Guoren He; Suxia Yuan; Jun Ming

doi:10.7717/peerj.4424

Tobacco rattle virus-induced PHYTOENE DESATURASE (PDS) and Mg-chelatase H subunit (ChlH) gene silencing in Solanum pseudocapsicum L

PeerJ. 2018 Mar 20:6:e4424. doi: 10.7717/peerj.4424. eCollection 2018.

Authors

Hua Xu^{1

2}, Leifeng Xu¹, Panpan Yang^{1

3}, Yuwei Cao¹, Yuchao Tang¹, Guoren He¹, Suxia Yuan¹, Jun Ming¹

Affiliations

¹ Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing, China.
² College of Life and Environmental Science, GanNan Normal University, Ganzhou, China.
³ College of Landscape Architecture, Nanjing Forestry University, Nanjing, China.

Abstract

Virus-induced gene silencing (VIGS) is an attractive tool for determining gene function in plants. The present study constitutes the first application of VIGS in S. pseudocapsicum, which has great ornamental and pharmaceutical value, using tobacco rattle virus (TRV) vectors. Two marker genes, PHYTOENE DESATURASE (PDS) and Mg-chelatase H subunit (ChlH), were used to test the VIGS system in S. pseudocapsicum. The photobleaching and yellow-leaf phenotypes of the silenced plants were shown to significantly correlate with the down-regulation of endogenous SpPDS and SpChlH, respectively (P ≤ 0.05). Moreover, the parameters potentially affecting the efficiency of VIGS in S. pseudocapsicum, including the Agrobacterium strain and the inoculation method (leaf syringe-infiltration, sprout vacuum-infiltration and seed vacuum-infiltration), were compared. The optimized VIGS parameters were the leaf syringe-infiltration method, the Agrobacterium strain GV3101 and the growth of agro-inoculated plants at 25°. With these parameters, the silencing efficiency of SpPDS and SpChlH could reach approximately 50% in S. pseudocapsicum. Additionally, the suitability of various reference genes was screened by RT-qPCR using three candidate genes, and the results demonstrated that glyceraldehyde 3-phosphate dehydrogenase (GAPDH) can serve as a suitable reference for assessing the gene expression levels of VIGS systems in S. pseudocapsicum. The proven application of VIGS in S. pseudocapsicum and the characterization of a suitable reference gene in the present work will expedite the functional characterization of novel genes in S. pseudocapsicum.

Keywords: Gene expression; Mg-chelatase H subunit (ChlH); PHYTOENE DESATURASE (PDS); Reference genes; Solanum pseudocapsicum L.; TRV; VIGS.

Grants and funding

This study was supported by the National Natural Science Foundation of China (31272205, 31672196), the Fundamental Research Funds for Central Non-profit Scientific Institutions, and the Science and Technology Innovation Program of the Chinese Academy of Agricultural Sciences. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.