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Over the past several decades, gene therapy technologies have been developed for a diverse number of applications ranging from DNA-based vaccines to gene silencing with RNAi. While all are powerful tools, a common limitation for these technologies is the need for effective and safe delivery to target sites within the body. Such delivery vectors are necessary for retention of bioactivity and stability, while also providing a method of cellular and tissue uptake and distribution, which may require endosomal escape. Although, viral and lipid-based technologies have shown promise as nucleic acid delivery vectors, both have inherent issues such as cytoxicity, oncogenicity, and immunogenicity. Thus, the development of polymer-based non-viral vectors has been an area of great focus over the past decade. While many polymeric vectors have been developed for plasmid DNA (pDNA) delivery, very few have shown effective delivery of short interfering RNA (siRNA), a powerful tool for gene silencing via the RNA interference mechanism. Furthermore, very few prospective delivery vectors have shown versatility for the administration of siRNA through multiple routes of administration. The overall goal of this research was to develop a biocompatible non-viral delivery system for the delivery of plasmid DNA, siRNA, and contrast agents through the modification of the natural biopolymer chitosan. We have synthesized an imidazole modified chitosan (chitosan-IAA) by conjugation of imidazole acetic acid to chitosan. Extensive evaluation and characterization of the modified polymer demonstrates enhanced solubility and buffering capacity within the physiological and endosomal pHs, thus providing enhanced endosomal escape by exploiting the "proton sponge" effect. We have demonstrated effective in vitro gene expression and gene silencing with chitosan-IAA mediated delivery of pDNA and siRNA, respectively. Furthermore, we have demonstrated in vivo gene silencing by delivery of siRNA through both intranasal and intravenous routes of delivery with chitosan-IAA/siRNA nanocomplexes. We have also demonstrated delivery of contrast agents up to 45 nm in size through mucosal tissue following treatment with chitosan and no contrast agent modification in both human and animal tissue. In conclusion, we have successfully developed a versatile and highly effective delivery vector for both nucleic acids and contrast agents.