Molecular Characterization of a Stable Antisense Chalcone Synthase Phenotype in Strawberry (Fragaria × ananassa)

Abstract

An octaploid (Fragaria x ananassa cv. Calypso) genotype of strawberry was transformed with an antisense chalcone synthase (CHS) gene construct using a ripening related CHS cDNA from Fragaria x ananassa cv. Elsanta under the control of the constitutive CaMV 35S promoter via Agrobacterium tumefaciens. Out of 25 transgenic lines, nine lines showed a reduction in CHS mRNA accumulation of more than 50% as compared to the untransformed cv. Calypso control. The antisense CHS construct was found to be integrated into the genome, with a copy number ranging from one to four. The pigmentation of the fruit was only affected when less than 5% of the control CHS expression level was detected. A stable antisense phenotype over a period of 4 years was obtained in the primary transgenic lines at a rate of 1:20. As a consequence of the reduced activity of CHS, the levels of anthocyanins, flavonols, and proanthocyanidins were downregulated and precursors of the flavonoid pathway were shunted to the phenylpropanoid pathway leading to highly increased levels of cinnamoyl glucose (520% of control), caffeoyl glucose (816% of control), and feruloyl glucose (1092% of control) as well as p-coumaryl alcohol (363% of control) and p-coumaryl-1-acetate (1079% of control), which occur only as trace components in untransformed control fruits. These results demonstrate that the introduction of an antisense CHS construct in strawberry results in an unpredictable biochemical phenotype, thereby confirming that CHS function is an important regulatory point of substrate flow between the flavonoid and the phenylpropanoid pathways.

Caracterización de indicadores de la calidad del fruto en líneas de fresa transgénicas con genes silenciados que codifican para enzimas pectinolíticas

“…Estos resultados contrastan con los descritos previamente por Jiménez-Bermúdez et al (2002), quienes, mediante un ensayo de penetración, observaron mayor firmeza en frutos de esta línea que en el control. La ausencia de diferencias significativas en firmeza entre la línea APEL 21 y el control descrita en el presente trabajo podría sugerir una pérdida de efectividad del silenciamiento inducido por el antisentido del gen FaplC tras varias rondas de multiplicación vegetativa del material, como se ha descrito previamente en otras líneas transgénicas de fresa (Lunkenbein et al, 2006). Sería necesario determinar nuevamente la expresión del gen FaplC en frutos de esta línea para confirmar esta hipótesis.…”

Acylphloroglucinol biosynthesis in strawberry fruit

“…Both have close homology to FaCHS (accession no. AI795154.1), which is highly expressed in strawberry fruit (Lunkenbein et al, 2006). Fig.…”

“…5) and fruits of a stable transgenic FaCHS antisense line (Lunkenbein et al, 2006) were performed. Fruits infiltrated with the control vector ripened normally to red, whereas fruits agroinfiltrated with plasmid binary (pBI)-CHSi became only slightly red (orange) colored (Fig.…”

“…S2). A previously reported chalcone synthase gene from strawberry CHS (Lunkenbein et al, 2006), whose product catalyzes one of the first steps in the flavonoid pathway, was chosen as a positive reporter gene (Vogt, 2010). Downregulation of CHS transcript levels rapidly leads to lower levels of pigmentation in Fragaria spp.…”

Flavonoid Accumulation Behavior in Response to the Abiotic Stress: Can a Uniform Mechanism Be Illustrated for All Plants?

“…Also Fragaria spp. has been studied for their genes and enzymes ( [25][26][27][28][29][30]; cited by [24]). In order to determine which pathway is preferred for biosynthesis secondary metabolites under abiotic stress factors, the expression of protein or enzymes associated with synthesis of secondary metabolites in the pathways should be determined and then compared with the control group-not stressed group.…”