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Carbon dots (CDs) have become the focus of many studies due to their outstanding optical properties and good biocompatibility. We investigated their potential application to produce a smart and highly efficient yet nontoxic nanovector for gene delivery. This was achieved by conjugating PEI 1.8k -functionalized CDs (synthesized by one-step microwave-assisted pyrolysis) with arginine-disulfide linkers to produce CD-PEI 1.8k -Arg nanoparticles. This nanovector could deliver p-CRISPR (9.3 kb) into different types of cell lines with higher efficiency compared to native PEI 1.8k or PEI 25k . CD-PEI 1.8k -Arg also maintained its outstanding transfection efficiency at a high serum concentration and low p-CRISPR dose, compared to PEI 25k, which was ineffective under those conditions. Additionally, CD-PEI 1.8k -Arg could knock out the GFP gene with great efficiency by delivering the required components of CRISPR/Cas9, including a plasmid encoding Cas9, sgRNA targeting GFP, and Cas9/sgRNA ribonucleoproteins (RNPs) into the HEK 293T-GFP cells. Moreover, the nanoparticles showed potential for the local delivery of p-CRISPR into brain tissue. The remarkable properties of CD-PEI 1.8k -Arg could enable the development of a safe, highly efficient gene-delivery nanovector for the treatment of various diseases in the near future.