Abstract
BackgroundThere are many varieties of carotenoids in pepper fruits. Capsanthin is a red carotenoid that gives mature pepper fruits their red color. The red color in pepper fruits is regulated mainly by the genes capsanthin/capsorubin synthase(Ccs), phytoene synthase(Psy), lycopene-β-cyclase(Lcyb) and β-carotene hydroxylase(Crtz). There has been very limited research work related to the development and change in the red color during fruit formation and when a certain gene or several genes are deleted. In this paper, we constructed viral vectors, using the tobacco rattle virus (TRV), to carry the target gene to infect detached pepper fruits, and observed the fruits’ color change. We used real-time quantitative PCR to analyze the gene silencing efficiency. At the same time, HPLC was used to determine the content of capsanthin and carotenoids that are associated with capsanthin synthesis when key genes in the pepper fruits were silenced.ResultsThese genes (Ccs, Psy, Lcyb and Crtz) were individually silenced through virus induced gene silencing (VIGS) technology, and pepper fruits from red fruit cultivars showed an orange or yellow color. When several genes were silenced simultaneously, the fruit also did not show the normal red color. Gene expression analysis by real-time quantitative PCR showed 70-80% efficiency of target gene silencing when using the VIGS method. HPLC analysis showed that the contents of carotenoids associated with capsanthin synthesis (e.g. β-carotene, β-cryptoxanthin or zeaxanthin) were decreased in varying degrees when silencing a gene or several genes together, however, the content of capsanthin reduced significantly. The synthesis of capsanthin was influenced either directly or indirectly when any key gene was silenced. The influence of the target genes on color changes in pepper fruits was confirmed via the targeted silencing of them.ConclusionsVIGS was a good method to study the molecular mechanism of pepper fruit color formation. By using virus induced gene silencing technology, capsanthin synthesis genes in pepper fruits were silenced individually or simultaneously, and pepper fruit color changes were observed. This provides a platform to further explore the molecular mechanism of pepper fruit color formation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-014-0314-3) contains supplementary material, which is available to authorized users.
Highlights
There are many varieties of carotenoids in pepper fruits
Effects of certain genes being silenced on pepper fruit color The capsanthin-capsorubin synthase (Ccs) gene carried by the tobacco rattle virus (TRV) viral vector was injected into detached fruits of Capsicum annuum cv
The color of the fruits injected with the TRV vector carrying the Ccs gene was from green to yellow (Figure 1d), while the control fruits were green to red color (Figure 1b)
Summary
Capsanthin is a red carotenoid that gives mature pepper fruits their red color. We constructed viral vectors, using the tobacco rattle virus (TRV), to carry the target gene to infect detached pepper fruits, and observed the fruits’ color change. HPLC was used to determine the content of capsanthin and carotenoids that are associated with capsanthin synthesis when key genes in the pepper fruits were silenced. Normal particle-producing isolates (called M-type) have two species of genomic RNA, i.e. RNA1 and RNA2. These are readily transmitted by inoculation with sap, and by nematodes in the family trichodoridae. The TRV vector induces very mild symptoms, infects large areas of adjacent cells and silences gene expression in growing points. Proteins encoded by RNA1 are sufficient for replication and movement within the host plant, while proteins encoded by RNA2 allow virion formation and nematode-mediated transmission between plants [1]
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