Overexpression and VIGS system for functional gene validation in oriental melon (Cucumis melo var. makuwa Makino)
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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.
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.
KeywordsTransient overexpression Virus-induced gene silencing (VIGS) Sprout absorption Lipoxygenases CmLOX10
Apple latent spherical virus
Bean pod mottle virus
Barley stripe mosaic virus
Cucumber mosaic virus
Green fluorescent protein
Potato virus X
Quantitative real-time PCR
Reactive oxygen species
Tobacco rattle virus
Virus-induced gene silencing
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).
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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