The use of recombinant DNA constructs has long been almost universal in biological research. Although retrovirus vectors are not as widely used, their utility for producing efficient ectopic or over-expression of proteins in primary cultured cells or embryos that has become more widely recognized in recent years. Despite this, not all laboratories doing musculoskeletal research have the capability to use their techniques to their fullest potential. Some laboratories have minimal experience in carrying out subcloning projects to make expression vectors, promoter- reporter constructs, or gene targeting vectors. These laboratories must try to collaborate with other laboratories, and this may not always be possible. Other laboratories have the capability to carry out subcloning projects but do not have the ability to generate retrovirus vectors in a cost efficient manner. Thus, they are unable to address important questions in the most relevant cell types, such as primary cell cultures or embryos, and most use transfected immortalized cell lines that often do not display the full range of the normal in vivo phenotype. Even laboratories that have the capability to make retrovirus vectors can benefit from assistance of personnel who are highly experienced in production of these vectors. The Vector Core will provide the services of technical personnel who will be highly trained in the production of DNA constructs and retrovirus vectors. In addition, Dr. Lichtler and the Core staff will work with investigators to design the most appropriate vector for the experimental purpose. The Core will maintain plasmid stocks of standard and skeletal cell type specific expression vectors, constructs needed to make gene targeting vectors and retrovirus vectors. Standardized, high titer stocks of retrovirus vectors expressing mak4er genes such as beta-galactosidase and green fluorescent protein will also be maintained to allow rapid evaluation of the suitability of a particular vector type for a proposed experiment. Self-inactivating retrovirus vectors containing skeletal type specific promoters will be developed and maintained for use as cell type specific markers in primary cell cultures. A fee structure will be developed to ensure that experiments are well thought out, and to help maintain the Core budget, but not prevent investigators from utilizing the Core. Thus, the Vector Core will facilitate the research of the members of the Core by providing advanced vectors which they otherwise would have difficulty in obtaining.
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