Establishment of protoplasts transient expression system in Pinellia ternata (Thunb.) Breit

In this study, we established an efficient and rapid transient expression system in the protoplasts of Pinellia ternata (Thunb.) Breit. (P. ternata). The protoplasts of P. ternata were prepared from plant leaves as the source material by digesting them with the combination of 20 g·l−1 cellulase and 15 g·l−1 macerozyme for 6 h. Based on the screening of PEG concentration, the conditions for PEG-mediated protoplast transformation were improved, and the highest transformation efficiency was found for 40% PEG 4000. Furthermore, we used the subcellular protein localization technique in P. ternata protoplasts to allow further validation of transient expression system. We present the method that can be applicable for studying both gene verification and expression in P. ternata protoplasts, thus allowing for engineering the improved varieties of P. ternata through molecular plant breeding techniques. This method can also be widely applicable for analyzing protein interactions, detecting promoter activity, for somatic cell fusion in plant breeding, as well as for other related studies.