Recombinant DNR Probe Technology in the Search For the Multiple Endocrine Neoplasia Type 2B Gene

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Drabik, Christopher E.
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Multiple Endocrine Neoplasia (MEN) type 2A and 2B are related diseases showing phenotypic overlap in the kinds of tumor involvement of each disease. Recently, by linkage to DNA markers, Multiple Endocrine Neoplasia type 2A (MEN 2A) has been localized to the pericentric region of chromosome 10 (Simpson and Kidd, 1987 and Mathews 1987). Because of phenotype overlap and consistent tumor involvement, it has been proposed that MEN type 2A and 2B should consist of overlapping genes causing the various syndromes of the disease (Jackson and Van Dyke et al. 1987). No evidence to date has been given showing that MEN 2A and 2B are linked. Isolating and characterizing either of the two loci could lead to a further understanding of the genetic and molecular aspects of cancer. Also, isolating markers near either of the genes could lead to better prediction of the diseases. Although, MEN 2A has been localized by probes to chromosome 10, the recombination frequency between the probes and MEN 2A is sufficiently large enough for prediction of MEN 2A to be ruled out. The generation of probes from a recombinant DNA library and the use of recombinant DNA probes for genetic linkage analysis in the search for the Men 2B gene locus was explored in this study. R cosmid library consisting of fragments of DNR from chromosome ten inserted into the vector pCos 2 and then cloned into E. coli was characterized using restriction enzymes. The recombinant cosmid clones were screened for unique sequences by hybridization to total human genomic DNA and then screened once more against DNR from chromosome ten only. 32 probes specific for chromosome 10 were generated from 250 cosmids. The previously localized chromosome ten probes D 1 OZ 1, D10S5 and RBP3 were analyzed for linkage to the Men 2B gene locus in 8 families affected with Men 2B. DNR from every available individual was digested with several restriction enzymes and the resulting fragments hybridized to D 1 OZ 1, 01 OS5 and RBP3. Families with informative polymorphism for the restriction enzymes were used to determine whether linkage to the probes existed. Lod scores were compiled for the Men 2B locus verse the loci represented by the probes and feasability of linkage to the probe loci determined. D 1 OZ 1, a probe consisting of a sequence of DNR from the centromere of chromosome 1 0, was found to be linked to MEN 2B. Linkage analysis of D 1 OS5 and RBP3 vs. MEN 2B was inconclusive.
42 p.
Kalamazoo College
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