DNA–Carbon Nanotube Binding Mode Determines the Efficiency of Carbon Nanotube-Mediated DNA Delivery to Intact Plants

Abstract

Efficient delivery of DNA, RNA, and genome engineering machinery to plant cells will enable efforts to genetically modify plants for global food security, sustainable energy production, synthetic biology applications, and climate change resilience. For the delivery of functional genetic units into plant cells, nanoparticles, particularly carbon nanotubes (CNTs), have attracted considerable interest. Although some success has been achieved using CNT-based approaches, the efficiency and practicality of the method for genome editing applications remain elusive. This is partly due to insufficient knowledge about the mechanisms of CNT-mediated delivery and expression of CNT-condensed DNA in plants. Here, we characterize the transcription and transformation efficiency of DNA deposited on CNTs coated with positively charged polymers by applying multiple experimental settings and reporter systems controlling the delivery and expression of DNA in plants. We found that the formation of partially condensed DNA on the CNT surface is a prerequisite for transfection and expression. In addition, we show that DNA irreversibly binds to the CNT and does not detach completely from the CNT surface. These results, together with an in vitro transcription assay, suggest that only the partially condensed part of the DNA is accessible to the cellular transcription machinery. Thus, the overall transcription and translation efficiency remains low, in particular for the large DNA units that are required for genome editing applications. Understanding the underlying mechanisms and limitations of CNT-mediated delivery of DNA through the plant cell wall is of considerable importance in guiding efforts to design nanomaterials for efficient transformation, agricultural trait engineering, and synthetic biology applications.

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