A major component of this Center is to use genetic manipulation of the mouse brain to unravel the molecular basis of learning and memory. One of the approaches here is to generate strains of mice in which a gene of interest has been eliminated in specific neuronal populations by use of the Cre recombinase. A prerequisite for this strategy is the availability of promoters that target Cre expression to specific populations of neurons in brain regions of interest. The purpose of this core is to identify appropriate genes and generate the targeted Cre mice. The PI proposes to isolate brain region specific transcripts using a combined subtractive hybridization-array screening strategy followed by in situ hybridization. This approach is based on existing commercial methods which require modification in most cases. The applicant is aware of this and has made substantial progress in optimizing the subtraction methodology. An alternative strategy, which the PI may well have considered, is to use in situ hybridization as the secondary screen (rather than using array technology). However, the proximity of the MIT Genome Center, which has extensive expertise in robotic screening methods, also favors the array approach. Following the isolation of transcripts, BACS will be identified for the clones having the requisite neuronal specificity and these used to generate BAC-Cre constructs. The use of BACs to construct transgenic mice has gained popularity and is probably the best approach available (assuming the candidate gene is represented in the commercial BAC libraries). The BAC fusion constructs will be used to make lines of transgenic mice that express Cre recombinase in specific populations of neocortical neurons. Subsequently, these mice will be crossed with lines of mice that harbor a gene of interest flanked by loxP sites.
Showing the most recent 10 out of 32 publications
