Factors affecting transformation of cell cultures from three dicotyledonous pigment-producing species using microprojectile bombardment

Abstract

Gene transfer methods were established for cell suspension cultures of sweet potato (Ipomoea batatas), ohelo (Vaccinium pahalae) and carrot (Daucus carota, two lines) using micro-projectile bombardment. Several parameters were studied (particle size/type, helium pressure, stage height, DNA concentration, pre-culture period) to determine which significantly affected transformation efficiency. All the physical parameters influenced transient gene expression, with particle size and type having the greatest effect. Cell culture age also affected transformation efficiency in all cell lines. Nuclear DNA conformation (relaxation) as measured by flow cytometry showed no change associated with culture age or transformation efficiency. Agrobacterium-based methods were also tested and in most experiments produced no GUS-expressing loci. Microprojectile bombardment will now be used to study anthocyanin biosynthesis in cell cultures as an alternative source of natural food colourants.