Establishment of an efficient genetic transformation method in Dunaliella tertiolecta mediated by Agrobacterium tumefaciens

Journal of Microbiological Methods
Claudia D Norzagaray-ValenzuelaAngel Valdez-Ortiz

Abstract

Microalgae are photosynthetic microorganisms widely used for the production of highly valued compounds, and recently they have been shown to be promising as a system for the heterologous expression of proteins. Several transformation methods have been successfully developed, from which the Agrobacterium tumefaciens-mediated method remains the most promising. However, microalgae transformation efficiency by A. tumefaciens is shown to vary depending on several transformation conditions. The present study aimed to establish an efficient genetic transformation system in the green microalgae Dunaliella tertiolecta using the A. tumefaciens method. The parameters assessed were the infection medium, the concentration of the A. tumefaciens and co-culture time. As a preliminary screening, the expression of the gusA gene and the viability of transformed cells were evaluated and used to calculate a novel parameter called Transformation Efficiency Index (TEI). The statistical analysis of TEI values showed five treatments with the highest gusA gene expression. To ensure stable transformation, transformed colonies were cultured on selective medium using hygromycin B and the DNA of resistant colonies were extracted after five subcultures and m...Continue Reading

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