Efficient leaf mesophyll protoplast isolation and transient gene expression in bermudagrasses (Cynodon spp.)

Bermudagrasses (Cynodon spp.) are important warm-season turfgrass species with multiple applications. Although physiological and high-throughput omics studies were conducted to explore the growth, development and stress responses of bermudagrasses, the molecular mechanism how their turf traits form remain unclear due to difficulties in genetic and transgenic manipulations. In this study, we reported an optimized enzymatic digestion method to stably isolate leaf mesophyll protoplasts from different bermudagrass cultivars. Furthermore, transient expression of two bermudagrass proteins, TEOSINTE BRANCHED1 and phytoene desaturase with different subcellular locations, were successfully observed using the isolated protoplasts. These results suggested that the leaf mesophyll protoplast system could be utilized in gene functional studies of bermudagrasses. As we known, this is the first systematic report of protoplast isolation and application in warm-season turfgrasses, especially Cynodon spp.