Main factors affecting the genetic transformation of chrysanthemum var. Micromargara

Abstract

Factors influencing Agrobacterium tumefaciens-mediated genetic transformation of plants have been widely reported: type and concentration of antibiotic, co-culture period, concentration of bacteria, concentration of acetosyringone, and of course, the type and age of the explants, as well as temperature conditions. However, it is not yet understood how these factors interact and how they affect the efficiency of the final transformation. The aim of this work was to evaluate the interaction of the three main factors affecting the transformation of chrysanthemum (Dendranthema grandiflora) var. Micromargara. A 2 3 factorial design was used with central points, in which three concentrations of A. tumefaciens (1.0, 1.5. and 2.0 of D.O. 600 ), three concentrations of acetosyringone (50 μM, 75 μM and 100 μM), and three co-culture periods (1, 2 and 3 days) were evaluated. The transformation was verified by GUS staining and by means of RT-PCR amplification and sequencing of fragment of the genes uidA, aph3 II and act-cr. The results show that, among the factors evaluated, only the concentration of A. tumefaciens presented a statistically significant effect (p< 0.05) on transformation efficiency, without interaction with the factors of acetosyringone concentration and co-culture period. It is concluded therefore that an A. tumefaciens concentration of D.O 600 : 2.0 is determinant for greater transformation efficiency in chrysanthemum var. Micromargara.