Viral vectors are the most efficient tools currently available for genetic modification of mammalian cells in vitro and in vivo. However, because protocols for the incorporation of DNA of interest into viral particles are rather lengthy and cumbersome, these vectors have not been extensively used in the area of cell biology. In the area of genetic therapy these vectors remain unchallenged tools, but they are expensive to produce, suffer from the inability to target only chosen types of cells and their application could be associated with substantial health risks. On the contrary, bacteriophage lambda-derived vectors are easy and safe to use. They have a substantial capacity and could be reproducibly obtained in high quantities. For years bacteriophage lambda-based vectors have been used as versatile tools for the delivery of foreign DNA into bacterial, but not into mammalian cells. The purpose of the current proposal is to identify ways to convert bacteriophage lambda from being a general toot in molecular biology to a general toot for cell biology. We intend to prove that provided with means to recognize specific receptors on the surface of mammalian cells and penetrate through the cell membrane, bacteriophage lambda particles will be able to deliver extended DNA sequences into mammalian cells. During Phase I proof of the concept will be achieved through the delivery of DNA capable of directing the synthesis of green fluorescent protein into mammalian cells. During Phase II efforts will be directed towards optimization of such important processes as receptor recognition, membrane penetration, release of DNA from a phage capsid and transport of DNA into the cell nucleus. As a result of these efforts we expect to create a new tool that will allow for efficient shuttling of DNA constructs (including combinatorial libraries) from E. coil into mammalian cells. Also, we expect to lay a foundation for construction of customized bacteriophage-based DNA delivery systems suitable for genetic therapy applications.
