A callus-derived regeneration and Agrobacterium -mediated gene transformation developed for bilberry, Vaccinium myrtillus

Vaccinium myrtillus, known as bilberry or European blueberry, is highly valued by consumers for its berries that are rich in health-promoting compounds. However, bilberry production remains largely wild, as the low fruit yield and soft berry characteristics impede its domestication and commercialization. A robust gene transformation system, which is currently lacking in bilberry, can be used to verify gene functions, as well as to accelerate the breeding process. In this study, we propagated bilberries in laboratory conditions and established a callus-derived regeneration method. We further performed Agrobacterium -mediated gene transformation on bilberry. With step-wise optimization, the transformation efficiency reached 4% using newly developed ternary vector systems in conjunction with sonication treatment and vacuum infiltration. We successfully transformed the MYBA1 transcription factor, which promotes anthocyanin synthesis, into the bilberry genome. Consistent expression of MYBA1 resulted in observable anthocyanin accumulation in bilberry callus. Overall, a highly efficient gene transformation method was established in bilberry, which will benefit future gene function studies and bioreactor research on bilberry.