Optimization of a virus-induced gene silencing system and functional elucidation of MaBAM9b in postharvest banana fruits

Abstract

<jats:title>Abstract</jats:title> <jats:p><jats:bold>Background</jats:bold>: The genetic basis of metabolic pathways that operate during fruit ripening needs to be understood before the nutritional value of the banana can be improved. The banana is a typical starch conversion fruit, and β-amylase is a key enzyme that may play an important role in starch degradation during the ripening process. <jats:italic>Musa acuminata β-amylase 9b </jats:italic>(<jats:italic>MaBAM9b</jats:italic>) is closely related to starch degradation. However, its exact function in starch degradation has not been demonstrated in banana. Stable genetic transformation to identify gene function is time- and energy-consuming. Thus, an efficient and rapid method is needed for functional identification. Virus-induced gene silencing (VIGS) is a reverse-genetics method based on RNA-mediated antiviral plant defense that has been used to rapidly identify gene functions in plants. The aim of this study is to optimize a VIGS system and functional elucidation of <jats:italic>MaBAM9b</jats:italic> in postharvest banana fruits. <jats:bold>Results</jats:bold>: Using 0.5% iodine-potassium-iodide (I<jats:sub>2</jats:sub>-KI) staining for 150 s, we found that 1:3 TRV1:TRV2-<jats:italic>MaBAM9b </jats:italic>cultivated at 30 mmHg for 30 s to an optical density (OD) of 0.8 at 600 nm, and kept on Murashige &amp; Skoog (MS) media for 5 days produced the best silencing results. Under these conditions, the total starch content was greatly increased, whereas the β-amylase activity, soluble sugar content, and expression of endogenous<jats:italic> MaBAM9b</jats:italic> greatly decreased. <jats:bold>Conclusions: </jats:bold>The system described here is particularly useful for studying genes and networks involved in starch conversion in fruit, which alone would not produce a visual phenotype. This system will provide a platform for functional genomics and fruit quality improvement in the banana.</jats:p>