title: Functional analysis of soybean genes involved in flavonoid biosynthesis by virus-induced gene silencing.
abstract: Virus-induced gene silencing (VIGS) is a powerful tool for functional analysis of genes in plants. A wide-host-range VIGS vector, which was developed based on the Cucumber mosaic virus (CMV), was tested for its ability to silence endogenous genes involved in flavonoid biosynthesis in soybean. Symptomless infection was established using a pseudorecombinant virus, which enabled detection of specific changes in metabolite content by VIGS. It has been demonstrated that the yellow seed coat phenotype of various cultivated soybean lines that lack anthocyanin pigmentation is induced by natural degradation of chalcone synthase (CHS) mRNA. When soybean plants with brown seed coats were infected with a virus that contains the CHS gene sequence, the colour of the seed coats changed to yellow, which indicates that the naturally occurring RNA silencing is reproduced by VIGS. In addition, CHS VIGS consequently led to a decrease in isoflavone content in seeds. VIGS was also tested on the putative flavonoid 3'-hydroxylase (F3'H) gene in the pathway. This experiment resulted in a decrease in the content of quercetin relative to kaempferol in the upper leaves after viral infection, which suggests that the putative gene actually encodes the F3'H protein. In both experiments, a marked decrease in the target mRNA and accumulation of short interfering RNAs were detected, indicating that sequence-specific mRNA degradation was induced. The present report is a successful demonstration of the application of VIGS for genes involved in flavonoid biosynthesis in plants; the CMV-based VIGS system provides an efficient tool for functional analysis of soybean genes.
authors.affiliation: Research Faculty of Agriculture, Hokkaido University, Sapporo 060-8589, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Nagamatsu
authors.forename: Chikara
authors.initials: C
authors.lastname: Masuta
authors.forename: Mineo
authors.initials: M
authors.lastname: Senda
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Matsuura
authors.forename: Atsushi
authors.initials: A
authors.lastname: Kasai
authors.forename: Jin-Sung
authors.initials: JS
authors.lastname: Hong
authors.forename: Keisuke
authors.initials: K
authors.lastname: Kitamura
authors.forename: Jun
authors.initials: J
authors.lastname: Abe
authors.forename: Akira
authors.initials: A
authors.lastname: Kanazawa
mesh_terms: D000217:Acyltransferases / Q000235:genetics* / Q000378:metabolism; D017799:Cucumovirus; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism; D015536:Down-Regulation; D005419:Flavonoids / Q000096:biosynthesis*; D044948:Flavonols / Q000378:metabolism; D020868:Gene Silencing; D005818:Genetic Engineering; D005822:Genetic Vectors; D007529:Isoflavones / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism; D010860:Pigments, Biological / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D012639:Seeds / Q000378:metabolism; D013025:Glycine max / Q000235:genetics* / Q000378:metabolism / Q000821:virology
chemical_list: D005419:Flavonoids; D044948:Flavonols; D007529:Isoflavones; D010860:Pigments, Biological; D012333:RNA, Messenger; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; C038888:flavonoid 3'-hydroxylase; D000217:Acyltransferases; C020141:flavanone synthetase