A centrifugation-assisted peptide-mediated gene transfer method for high-throughput analyses

Abstract

A peptide-mediated DNA delivery system for several plant species has been recently developed. This system uses ionic complexes of DNA and fusion peptides containing several domains, such as DNA-binding and cell-penetrating peptides. Although the peptide-DNA complexes are capable of penetrating into plant cells through the cell wall by mechanical pressure using a syringe, sample throughput is limited. Here, we describe a Centrifugation-Assisted Peptide-mediated gene Transfer (CAPT) method for improving sample throughput with reproducible gene transfer efficiency. We optimized the parameters of CAPT for transient gene transfer efficiency by using Nicotiana tabacum cotyledons as a model plant material. The optimal parameters for centrifugation were 10,000×g for 60 s. Furthermore, we successfully transferred the peptide–DNA complex into rice cotyledons using the optimized CAPT method. Thus, the CAPT method is superior to the previous syringe-mediated infiltration method in terms of sample throughput in multiple plant species.