Bulk segregated analysis, transcriptome, and functional analyses reveal the involvement of VcKCS19 and VcCER2 in regulating blueberry fruit wax accumulation

The blueberry ( Vaccinium L.) fruit is known for its light-blue appearance, attributed to its blue-black skin covered with a white waxy layer. This layer is crucial for the growth and storage of blueberry fruits. This study established an F 1 mapping population of 166 plants derived from ‘Chandler’ (wax-rich) and ‘Black Pearl’ (wax-poor). Bulk segregated analysis sequencing was used to identify the candidate region associated with cuticular wax, which was finally narrowed to a 1.85 Mb segment on chromosome 11, including ten candidate genes. Further transcriptome analyses and qRT-PCR revealed that the relative expression levels of five candidate genes ( VcMYB36 , VcDXS , VcCAMS1 , VcCER2, and VcKCS19 ) were different between ‘Chandler’ and ‘Black Pearl’. Combined with gene function annotation, we hypothesized that VcKCS19 and VcCER2 are key candidate genes for wax synthesis in blueberry, potentially accounting for the variations in wax content between ‘Chandler’ and ‘Black Pearl’. Transient overexpression and virus-induced gene silencing in blueberry fruit revealed that VcCER2 and VcKCS19 positively regulate wax accumulation in blueberry fruits. This study provided key insights into the molecular basis of wax synthesis in blueberry fruits, which provides new avenues for enhancing wax quality in genetic breeding programs.