title: A viral satellite DNA vector-induced transcriptional gene silencing via DNA methylation of gene promoter in Nicotiana benthamiana.
abstract: Virus-induced gene silencing (VIGS) has been widely used for plant functional genomics study at the post-transcriptional level using various DNA or RNA viral vectors. However, while virus-induced transcriptional gene silencing (VITGS) via DNA methylation of gene promoter was achieved using several plant RNA viral vectors, it has not yet been done using a satellite DNA viral vector. In this study, a viral satellite DNA associated with tomato yellow leaf curl China virus (TYLCCNV), which has been modified as a VIGS vector in previous research, was developed as a VITGS vector. Firstly, the viral satellite DNA VIGS vector was further optimized to a more convenient p1.7A+2mβ vector with high silencing efficiency of the phytoene desaturase (PDS) gene in Nicotiana benthamiana plants. Secondly, the constructed VITGS vector (TYLCCNV:35S), which carried a portion of the cauliflower mosaic virus 35S promoter, could successfully induce heritable transcriptional gene silencing (TGS) of the green fluorescent protein (GFP) gene in the 35S-GFP transgenic N. benthamiana line 16c plants. Moreover, bisulfite sequencing results revealed higher methylated cytosine residues at CG, CHG and CHH sites of the 35S promoter sequence in TYLCCNV:35S-inoculated plants than in TYLCCNV-inoculated line 16c plants (control). Overall, these results demonstrated that the viral satellite DNA vector could be used as an effective VITGS vector to study DNA methylation in plant genomes.
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: zhengju2016@126.com.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Ju
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: lwang2015@126.com.
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: caody2012@126.com.
authors.forename: Dongyan
authors.initials: D
authors.lastname: Cao
authors.affiliation: National Engineering Research Center for Vegetables, Beijing Academy of Agriculture and Forestry Sciences, Beijing 100097, China. Electronic address: zuojinhua@126.com.
authors.forename: Jinhua
authors.initials: J
authors.lastname: Zuo
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: hlzhu@cau.edu.cn.
authors.forename: Hongliang
authors.initials: H
authors.lastname: Zhu
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: daqifu@126.com.
authors.forename: Daqi
authors.initials: D
authors.lastname: Fu
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: lyb@cau.edu.cn.
authors.forename: Yunbo
authors.initials: Y
authors.lastname: Luo
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China. Electronic address: zbz@cau.edu.cn.
authors.forename: Benzhong
authors.initials: B
authors.lastname: Zhu
mesh_terms: D019175:DNA Methylation; D004267:DNA Viruses / Q000235:genetics; D004276:DNA, Satellite; D004279:DNA, Viral; D015870:Gene Expression; D023061:Gene Order; D020868:Gene Silencing; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics*; D054884:Host-Pathogen Interactions / Q000235:genetics; D010641:Phenotype; D010935:Plant Diseases / Q000235:genetics / Q000821:virology; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D014026:Nicotiana / Q000235:genetics* / Q000821:virology*
chemical_list: D004276:DNA, Satellite; D004279:DNA, Viral
keywords: Bisulfite sequencing; DNA methylation; Green fluorescent protein; Satellite DNA virus; Tomato yellow leaf curl China virus; Virus-induced transcriptional gene silencing