title: An efficient virus-induced gene silencing vector for maize functional genomics research.
abstract: Maize is a major crop whose rich genetic diversity provides an advanced resource for genetic research. However, a tool for rapid transient gene function analysis in maize that may be utilized in most maize cultivars has been lacking, resulting in reliance on time-consuming stable transformation and mutation studies to obtain answers. We developed an efficient virus-induced gene silencing (VIGS) vector for maize based on a naturally maize-infecting cucumber mosaic virus (CMV) strain, ZMBJ-CMV. An infectious clone of ZMBJ-CMV was constructed, and a vascular puncture inoculation method utilizing Agrobacterium was optimized to improve its utility for CMV infection of maize. ZMBJ-CMV was then modified to function as a VIGS vector. The ZMBJ-CMV vector induced mild to moderate symptoms in many maize lines, making it useful for gene function studies in critically important maize cultivars, such as the sequenced reference inbred line B73. Using this CMV VIGS system, expression of two endogenous genes, ZmPDS and ZmIspH, was found to be decreased by 75% and 78%, respectively, compared with non-silenced tissue. Inserts with lengths of 100-300 bp produced the most complete transcriptional and visual silencing phenotypes. Moreover, genes related to autophagy, ZmATG3 and ZmATG8a, were also silenced, and it was found that they function in leaf starch degradation. These results indicate that our ZMBJ-CMV VIGS vector provides a tool for rapid and efficient gene function studies in maize.
authors.affiliation: State Key Laboratory for Agro-Biotechnology and Department of Plant Pathology, China Agricultural University, Beijing, 100193, China.
authors.forename: Rong
authors.initials: R
authors.lastname: Wang
authors.affiliation: State Key Laboratory for Agro-Biotechnology and Department of Plant Pathology, China Agricultural University, Beijing, 100193, China.
authors.forename: Xinxin
authors.initials: X
authors.lastname: Yang
authors.affiliation: State Key Laboratory for Agro-Biotechnology and Department of Plant Pathology, China Agricultural University, Beijing, 100193, China.
authors.forename: Nian
authors.initials: N
authors.lastname: Wang
authors.affiliation: State Key Laboratory for Agro-Biotechnology and Department of Plant Pathology, China Agricultural University, Beijing, 100193, China.
authors.forename: Xuedong
authors.initials: X
authors.lastname: Liu
authors.affiliation: Plant Biology Division, The Samuel Roberts Noble Foundation Inc., 2510 Sam Noble Parkway, Ardmore, OK 73401, USA.
authors.forename: Richard S
authors.initials: RS
authors.lastname: Nelson
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, 12 South Zhongguancun Street, Beijing, 100081, China.
authors.forename: Weimin
authors.initials: W
authors.lastname: Li
authors.affiliation: State Key Laboratory for Agro-Biotechnology and Department of Plant Pathology, China Agricultural University, Beijing, 100193, China.
authors.forename: Zaifeng
authors.initials: Z
authors.lastname: Fan
authors.affiliation: State Key Laboratory for Agro-Biotechnology and Department of Plant Pathology, China Agricultural University, Beijing, 100193, China.
authors.forename: Tao
authors.initials: T
authors.lastname: Zhou
mesh_terms: D060054:Agrobacterium / Q000235:genetics; D001343:Autophagy; D001483:Base Sequence; D017799:Cucumovirus / Q000502:physiology*; D020868:Gene Silencing; D005822:Genetic Vectors / Q000235:genetics*; D023281:Genomics; D007178:Inbreeding; D018515:Plant Leaves / Q000235:genetics / Q000648:ultrastructure; D003313:Zea mays / Q000235:genetics* / Q000648:ultrastructure
keywords: autophagy; cucumber mosaic virus; functional genomics study; knockdown; maize inbred lines; vascular puncture inoculation