TEMPORAL EXPRESSION OF DNP-SPECIFIC V-GENES

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Previous work has established that the anti-dinitrophenyl (DNP) response in neonatal Balb/c mice is restricted to only a few idiotypes whereas the adult response is quite diverse. This behavior roughly parallels the overall expansion of the Balb/c immune repertoire: newborn mice express a total of about 10(4) different specificities (clonotypes) and this gradually increases to greater than 10(7) clonotypes in the adult. The aim of the proposed program is to characterize, at the DNA level, the temporal development of the anti-DNP response in Balb/c mice. In collaboration with Dr. Norman Klinman here at Scripps Clinic and Research Foundation, we will prepare hybridomas expressing distinct anti-DNP idiotypes and determine the temporal expression of these idiotypes using the Klinman splenic focus assay. Three hybridomas expressing common Balb/c neonatal clonotypes have already been constructed in Klinman's laboratory. Using recombinant DNA methods, we will determine the DNP-specific V-, D-, and J-gene segment sequences expressed in each of these hybridomas and the relative locations of these gene segments in Balb/c germline DNA. We will then try to understand the temporal expression of each gene product in terms of its sequence and flanking sequences, and its location in the germline genome. We will extend this work to other inbred mouse strains; this will allow us to test our interpretations of the Balb/c data as well as to probe the molecular basis for strain-dependent differences in the anti-DNP response.