Transformation of European Ash ( L.) Callus as a Starting Point for Understanding the Molecular Basis of Ash Dieback.

The population of European ash ( L.) is currently facing the risk of collapse, mainly due to ash dieback, a disease caused by a pathogenic fungus, . To facilitate studies into the molecular basis of ash dieback and design breeding strategies for a generation of resistant trees, it is necessary to develop tools enabling the study of gene function in . Despite this, a method for the genetic engineering of is still missing. Here, we report the first successful genetic transformation of callus and a selection process enabling the formation of stable transgenic callus lines. The protocol relies on the use of to transform callus tissue derived from embryos of . In our experiments, we used the β-glucuronidase (GUS) reporter system to demonstrate the transformation of callus cells and performed RT-PCR experiments to confirm the stable expression of the transgene. Since ash dieback threatens the long-term stability of many native populations, we hope that the transformation techniques described in this manuscript will facilitate rapid progress in uncovering the molecular basis of the disease and the validation of gene targets previously proposed to be linked to the resistance of trees to pathogenicity.