An efficient papaya leaf distortion mosaic potyvirus vector for virus-induced gene silencing in papaya

Decai Tuo,Yang Liu,Guangyuan Zhao,Peng Zhou,Xiukun Yang,Guopeng Zhu,Hongguang Cui,Xiaoying Li,Wentao Shen,Pu Yan,He Wang

doi:10.1038/s41438-021-00579-y

Abstract

Papaya (Carica papaya L.) is regarded as an excellent model for genomic studies of tropical trees because of its short generation time and its small genome that has been sequenced. However, functional genomic studies in papaya depend on laborious genetic transformations because no rapid tools exist for this species. Here, we developed a highly efficient virus-induced gene silencing (VIGS) vector for use in papaya by modifying an artificially attenuated infectious clone of papaya leaf distortion mosaic virus (PLDMV; genus: Potyvirus), PLDMV-E, into a stable Nimble Cloning (NC)-based PLDMV vector, pPLDMV-NC, in Escherichia coli. The target fragments for gene silencing can easily be cloned into pPLDMV-NC without multiple digestion and ligation steps. Using this PLDMV VIGS system, we silenced and characterized five endogenous genes in papaya, including two common VIGS marker genes, namely, phytoene desaturase, Mg-chelatase H subunit, putative GIBBERELLIN (GA)-INSENSITIVE DWARF1A and 1B encoding GA receptors; and the cytochrome P450 gene CYP83B1, which encodes a key enzyme involved in benzylglucosinolate biosynthesis. The results demonstrate that our newly developed PLDMV VIGS vector is a rapid and convenient tool for functional genomic studies in papaya.

Highlights

Papaya (Carica papaya L., Caricaceae) is an economically important fruit tree cultivated in tropical and subtropical regions[1]
To further assess the ability of the pPLDMV-Nimble Cloning (NC) virus-induced gene silencing (VIGS) vector to silence endogenous genes in papaya, we investigated the roles of two putative papaya GID1A (PaGID1A) and GID1B (PaGID1B) in the development of papaya plants
In this study, we report the development of papayainfecting potyvirus as a VIGS vector for RNA silencing of endogenous papaya genes

Summary

Introduction

Papaya (Carica papaya L., Caricaceae) is an economically important fruit tree cultivated in tropical and subtropical regions[1]. In many tropical countries, other parts of the papaya plant, including seeds, leaves, roots, and flowers, are valued for their use in traditional medicine[4]. Unripe (green) papaya fruit produces a Papaya is regarded as an excellent model system for genomic studies of both tropical trees and fruit trees because of its small diploid genome of 372 megabases (Mb) with nine pairs of chromosomes, its short generation time of 9–15 months, and its continual year-long flower and fruit production[2,6]. In 2008, a draft genome sequence of “SunUp” papaya, the first transgenic commercial papaya ringspot virus (PRSV)-resistant fruit crop, was obtained using a whole-genome shotgun approach and represented the fifth complete genome sequence of a flowering plant after Arabidopsis thaliana, Oryza sativa (rice), Populus trichocarpa (poplar), and Vitis vinifera (grape)[2]. The sequenced genome has facilitated investigations of the functions of many relatively poorly studied

Methods

Results

Conclusion