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Overexpression and VIGS system for functional gene validation in oriental melon (Cucumis melo var. makuwa Makino)

  • Jing Jing Liao
  • Cheng Hui Wang
  • Qiao Juan Xing
  • Yue Peng Li
  • Xian Feng Liu
  • Hong Yan QiEmail author
Original Article
  • 34 Downloads

Abstract

As one of the effective and powerful methods for gene function analysis in plants, transient overexpression and virus-induced gene silencing (VIGS) system were developed for oriental melon (Cucumis melo var. makuwa Makino). Here, the full-length of CmLOX10, the conserved domains of CmLOX10 and CmPDS were isolated from oriental melon. The vectors for overexpression of CmLOX10, and the VIGS vector of CmLOX10 and CmPDS were constructed and transformed into agrobacterium, respectively. Especially, the sprout absorption method was performed on oriental melon, and CmPDS was selected as a reporter gene in VIGS. The expression level of CmLOX10 significantly increased in the oriental melon with the CmLOX10 overexpression construct, and lipoxygenases (LOXs) contributed to the leaf cell death in the oriental melon. The expression levels of CmLOX10 and CmPDS were significantly decreased in plants with the VIGS construct, and there was albino phenotype in CmPDS silencing leaves. In this study, CmLOX10 gene was silenced using Tobacco rattle virus based VIGS system firstly. These results proved that the transient expression technology provides an excellent tool to explore gene function in oriental melon.

Key message

Transient over-expression and virus-induced gene silencing (VIGS) system could be performed successfully in oriental melon by leaf agro-infiltration and sprout absorption methods, respectively.

Keywords

Transient overexpression Virus-induced gene silencing (VIGS) Sprout absorption Lipoxygenases CmLOX10 

Abbreviations

ALSV

Apple latent spherical virus

AS

Acetosyringone

BPMV

Bean pod mottle virus

BSMV

Barley stripe mosaic virus

CMV

Cucumber mosaic virus

DAB

Diaminobenzidine

GFP

Green fluorescent protein

LB

Luria-Bertani

LOXs

Lipoxygenases

MDA

Malondialdehyde

MES

4-Morpholineethanesulfonic acid

PDS

Phytoene desaturase

PEG

Polyethylene glycol

PVX

Potato virus X

qRT-PCR

Quantitative real-time PCR

ROS

Reactive oxygen species

TBA

Thiobarbituric acid

TCA

Trichloroacetic acid

TRV

Tobacco rattle virus

VIGS

Virus-induced gene silencing

Notes

Acknowledgements

We are grateful to Dr Hao Jinghong (Beijing University of Agriculture, China) for kindly offering pTRV1 and pTRV2 vectors. This study has been supported by China Agriculture Research System (Grant No.: CARS-25), Shenyang Science and Technology Project (Grant No.: 17-143-3-00) and Liaoning Innovative Research Team in University (Grant No.: LZ2015025).

Author contributions

JJL and HYQ conceived and designed the experiments. JJL, CGW, QJX, YPL and XFL performed the experiments. JJL analyzed the data and wrote the manuscript, and HQ improved the manuscript.

Compliance with ethical standards

Conflict of interest

All authors declare that they have no conflict of interest.

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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • Jing Jing Liao
    • 1
    • 2
  • Cheng Hui Wang
    • 1
  • Qiao Juan Xing
    • 1
  • Yue Peng Li
    • 1
  • Xian Feng Liu
    • 1
  • Hong Yan Qi
    • 1
    Email author
  1. 1.College of Horticulture and Shenyang Agricultural University/Key Laboratory of Protected Horticulture, Ministry of Education/Northern National & Local Joint Engineering Research Center of Horticultural Facilities Design and Application TechnologyShenyangChina
  2. 2.Institute of Medicinal Plant and Chinese Academy of Medical Sciences, Peking Union Medical CollegeBeijingChina

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