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An effective transformation protocol for  Dunaliella , a β-carotene producer, was developed using the synergistic mechanism of D-glucose and Acetosyringone on three different  Agrobacterium  strains (EHA105, GV3101 and LBA4404). In the present study, we investigated the pre-induction of  Agrobacterium  strains harboring pMDC45 binary vector in TAP media at varying concentrations of D-glucose (5 mM, 10 mM, and 15mM) and 100 μM of Acetosyringone for co-cultivation. Induction of  Agrobacterium  strains with 10 mM D-glucose and 100 μM Acetosyringone showed higher rates of efficiency compared to other treatments. The presence of GFP and HPT transgenes as a measure of transformation efficiency from the transgenic lines were determined using fluorescent microscopy, PCR, and southern blot analyzes. Highest transformation rate was obtained with the  Agrobacterium  strain LBA4404 (181 ± 3.78 cfu per 10 6  cells) followed by GV3101 (128 ± 5.29 cfu per 10 6  cells) and EHA105 (61 ± 5.03 cfu per 10 6  cells). However, the  Agrobacterium  strain GV3101 exhibited more efficient single copy transgene (HPT) transfer into the genome of  D .  salina  than LBA4404. Therefore, future studies dealing with genetic modifications in  D .  salina  can utilize GV3101 as an optimal  Agrobacterium  strain for gene transfer.

Synergistic Action of D-Glucose and Acetosyringone on Agrobacterium Strains for Efficient Dunaliella Transformation