Molecular biology, which in a broad sense covers the manipulation of DNA and RNA, has become an integral part of most biological research programs. Research areas that utilize molecular biology techniques include cloning genes for expression of proteins in cell model systems, site-directed mutagenesis and tagging for cellular localization and functional studies, genotyping, RNA isolation and expression analysis, and the development of transgenic mice. The Molecular Biology Core was established at the University of North Carolina (UNC) Cystic Fibrosis Center in 1998 to provide molecular biology services and expertise to faculty who are often primarily trained in cell biology, physiology, or clinical sciences. The Core is currently an integral part of many of the ongoing projects described in the PPG providing expert guidance, technical support, and specialized equipment. All four projects within the PPG application will utilize Molecular Biology Core services. PPG investigators will require cloning of genes into retroviral, lentiviral and oocyte expression vectors for expression of a variety of tagged and/or modified proteins (CFTR, ENaC subunits, and nucleoside tranporters) in a variety of cell models, including well-differentiated human bronchial epithelial (HBE) (provided by Core C:Cell Culture). Secondly, RNA isolation and expression analysis services, including quantitative real-time PCR and development and testing of si/shRNA reagents, will be provided, along with more specialized services, such as RNA-sequencing and data analyses. Finally, the Molecular Biology Core will aid in the generation and utilization of new transgenic mouse models, i.e., Panxl and Slc17A9, to help identify important regulators of ainway surface liquid homeostasis. The Molecular Biology Core will effectively complement the expertise of project leaders and their staff and to allow them to focus on the experiments in their own personal areas of proficiency.
The PPG focuses on pulmonary liquid homeostasis, attacking the problem from multiple vantage points in various cells, lung compartments, model systems. In as much as molecular biology can contribute to the successful completion of the goals of the grant, the Molecular Biology Core is highly relevant to the overall PPG. In this era of defining molecular mechanisms and links between muliple protein functions in complex model systems, molecular biology aspects of research projects will continue to be central and highly relevant.
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