Optimisation of introducing foreign genes into egg cells and zygotes of wheat (Triticum aestivum L.) via microinjection

An expeditious and highly efficient technique of microinjection has been de veloped with the aim of introducing exogenous DNA into egg cells and zygote s of wheat. Using a mechanical-dissection method and a novel immobilisation approach enabled us to microinject around 15 egg cells of wheat per hour. Exposing the protoplasts to a high-frequency alternating-current field for immobilisation, a significantly higher transient expression rate of the inj ected genes (46% and 52% for egg cells and zygotes, respectively) could be achieved than reported thus far for plant protoplasts. Whether this high tr ansformation efficiency is due to the high-frequency electrical field appli ed for immobilising the protoplasts is not known. The transformation rate a ppeared to be a factor depending upon the time of egg cell isolation. Accor ding to the ultrastructural observations this seems to reflect a variation in competence of the egg cells during in situ development. In order to cond uct studies directed towards establishing the optimal time-window for DNA d elivery into the fertilised egg cell, the time course of DNA dynamics durin g zygotic development has been quantified via quantitative microspectrofluo rometry.