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

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

doi:10.7717/peerj.4424

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.

Highlights

Solanum pseudocapsicum L., which belongs to the genus Solanum and the family Solanaceae, possesses high ornamental value and is widely cultivated as an indoor ornament due to its bright red berries at maturity (Aliero, Grierson & Afolayan, 2006b)
The results showed that the following fragments were obtained: approximately 600-bp ACTIN and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) fragments, 200-bp UBQ fragment, 600-bp PHYTOENE DESATURASE (PDS) fragment and 500-bp chelatase H subunit (ChlH) fragment (Fig. 1)
A BLASTX search of the sequence results was conducted, and the search results showed that the ACTIN, GAPDH, UBQ, PDS and ChlH amplification fragments were derived from the corresponding genes

Summary

Introduction

Solanum pseudocapsicum L., which belongs to the genus Solanum and the family Solanaceae, possesses high ornamental value and is widely cultivated as an indoor ornament due to its bright red berries at maturity (Aliero, Grierson & Afolayan, 2006b). This plant has important pharmaceutical value, as reflected by its use for the treatment of boils, How to cite this article Xu et al (2018), Tobacco rattle virus-induced PHYTOENE DESATURASE (PDS) and Mg-chelatase H subunit (ChlH ) gene silencing in Solanum pseudocapsicum L. The development of techniques for identifying the functions of genes in S. pseudocapsicum is vital

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Discussion

Conclusion