Associate Professor Kumaran has a long-standing interest in developing improved gene delivery and expression systems in mammalian cells. Work in his lab has developed several innovative applications using natural microbial mechanisms. In the 1990s his research adapted the natural homologous recombination mechanism operating in E. coli to develop recombination-based technology for genetic manipulation of very large DNA - a task that was extremely difficult until then. Another research project engineered innocuous E. coli to invade mammalian cells to deliver and express human genes. More recently his team has assembled artificial chromosomes that can be used to maintain and achieve long-term expression of human genes. Together these innovations seek to contribute to the field of gene delivery for the correction of human genetic disease and have led to a number of patents in this area. Associate Professor Kumaran is a member of the School of Science’s Industry Advisory Board, which provides strategic advice to the overall direction of the School. My other role is to drive industry engagement and industry led research on Campus.

1. Recombineering: DNA engineering using homologous recombination

Recombineering permits genetic engineering of DNA using homologous recombination in E. coli. Because homologous recombination is not dependent on the presence of suitably placed restriction enzyme sites like conventional genetic engineering methods, recombineering permits more complex manipulations and is not limited by the size of the DNA. Using this technique, various changes can be added to very large (>100 kb) DNA sequences including i) point mutations, ii) deletions, iii) insertions, and iv) gene fusions to develop specific cell lines and animal models of human disease. My current research is directed to better understand the action of the recombineering enzymes on various DNA substrates and to advance improved applications using this system.

2. E. coli as a vector for gene delivery into mammalian cells

Intracellular bacteria such as Shigella, Yersinia, and Salmonella, have evolved the capability to enter mammalian cells by invasion to establish pathogenicity. My research has adapted a non-pathogenic E. coli strain to express the Y. pseudotuberculosis invasin gene to deliver high molecular weight DNA up to 200 kb in size into mammalian cells by invasion. A vector capable of delivering such large DNA can include complete genes together with their introns, exons, and regulatory regions to permit more accurate expression of a genetic locus. Efforts are ongoing to improve the gene delivery efficiency of this vector into various mammalian cell types.

3. Artificial chromosome research

Gene therapy requires development of improved vectors for long-term retention and accurate expression of transgenes in cells. Currently available vectors can deliver genes efficiently into cells but provide only short bursts of expression before they become silenced by the host cell. To overcome these limitations my research is developing artificial chromosome vectors. Artificial chromosomes will 1) enable long-term retention of the delivered transgenes by existing as independent chromosomes that segregate faithfully to daughter cells during cell division, 2) provide long-term gene expression by avoiding DNA integration into human chromosomes, which can cause transgene silencing, and 3) provide correct levels and duration of expression because they carry complete genes along with their surrounding sequences that function to provide regulation.

• BTH1802 (Fundamentals of biotechnology)