Isolation of leaf mesophyll protoplasts optimized by orthogonal design for transient gene expression in Catalpa bungei

•An efficient protocol for protoplasts isolation from Catalpa bungei leaves was presented.•The experimental parameters were optimized by orthogonal experiment.•The viability of the protoplasts was up to 90 %.•PEG-mediated transient transformation system in C. bungei protoplasts was verified.