Metabolically engineered anthocyanin-producing lime provides additional nutritional value and antioxidant potential to juice

Abstract

Anthocyanins are synthesized via the flavonoid pathway through a complex expression of several genes such as MYB transcription factors. Anthocyanins protect plants against biotic and abiotic stresses. Herein, we studied the effect of expression of MYB (VvmybA1 cloned from the red grape and Ruby cloned from ‘Moro’ blood orange) transcription factors in “Mexican” lime on juice quality and leaf pigments, leaf metabolites, and phytohormones. Anthocyanins, furanocoumarins, flavonoids, and hydroxycinnamates were analyzed with high-performance liquid chromatography–mass spectrometry, whereas chlorophylls, carotenoids, and xanthophylls were analyzed using HPLC coupled with photodiode array detector (PDA). The rest of metabolites were analyzed using gas chromatography–mass spectrometry. Overexpression of VvmybA1 and Ruby resulted in accumulation of anthocyanins in leaves, flowers, and fruits of the transgenic plants. However, the level of anthocyanins in Ruby plants was significantly lower than that in VvmybA1 plants. The level of anthocyanins and the gene expression of VvmybA1 and Ruby in young leaves were higher than mature leaves. On the other hand, the level of several furanocoumarins, and hydroxycinnamates decreased in mature VvmybA1 leaves, indicating a drainage of p-coumaric acid due to the induction of anthocyanins biosynthesis. The level of chlorophyll decreased in mature VvmybA1 leaves, whereas zeaxanthin level increased, indicated a photoprotection role for anthocyanins. Most of polar and volatile metabolites also decreased VvmybA1 leaves, indicating a decrease in the photosynthetic efficiency. Benzoic acid and salicylic acid increased, whereas auxins decreased. The level of abscisic acid was not affected by the overexpression of VvmybA1 and the plants showed normal growth and development. Overexpression of VvmybA1 highly increased the antioxidant activity of the transgenic juice and leaves, whereas overexpression of Ruby showed only a slight increase. The pH, °Brix value, and TA of the transgenic juice were not affected by the expression of VvmybA1 or Ruby.