A Noninvasive Delivery of DNA into Zygotes by AuNPs−PEI for Transgenic Mice

Graphical Plasmid DNA coheres to the surface of AuNPs through electrostatic adsorption of the PEI mixture with a certain proportion of high-molecular-weight (25,000) PEI (HMW PEI) and low-molecular-weight (1,800) PEI (LMW PEI). AuNPs−PEI−DNA shows a very low toxic impact to embryo development and excellent efficiency of delivering exogenous DNA into mouse early embryonic cells. A transgenic system of noninvasive cytosolic transport based on exogenous plasmid DNA (pIRES2−EGFP) loading on gold nanoparticles (AuNPs) coated with polyethyleneimine (PEI) was successfully constructed, which can remove the technical threshold and possible damage of microinjection to oocytes and embryo development. In this study, mouse zygotes were cultured in KSOM medium containing the complex of AuNPs−PEI−DNA (APD) until formation of blastocyts, and then transferred into pseudopregnant female mice for obtaining transgenic offspring. Fluorescence observations confirm the expression of GFP in early embryos; importantly, data from identifying experiments by genotyping, immunofluorescence and DNA sequencing show that the GFP sequence indeed integrates into the chromosome of offspring individuals and is expressed. Furthermore, TUNEL test and analysis on cell differentiation of blastocysts indicate an excellent safety of APD to the development of embryos. All these results suggested that AuNPs, being an alternative technique to traditional microinjection, can efficiently introduce biomolecules into early embryonic cells without the prerequisite of expensive micromanipulators and skilled technicians, which will greatly promote study of life sciences in various aspects.