Transcript characterization in the region of the mouse neuromuscular mutationmnd2.

The mouse mutation mnd2 provides an experimental model for studying human neuromuscular and neurodegenerative disease. In order to identify the mnd2 gene, a positional cloning approach was taken. Linkage analysis demonstrated that mnd2 maps to a 0.2 $\pm$ 0.1 cM nonrecombinant interval in the central region of mouse chromosome 6, a region corresponding to human chromosome 2p13. To span this region, a contig of seven BAC clones and seven P1 clones was generated. The physical size of the nonrecombinant interval was between 360 kb and 500 kb. Four previously known genes were mapped to the nonrecombinant region: dynactin 1, smooth muscle gamma-actin, rhotekin, and p62Dok. Five novel genes in the region were identified by exon amplification, cross-species conservation, direct genomic sequencing, and mouse/human comparative sequence analysis. These are: a new member of the semaphorin gene family, semaphorin M; a new member of the WD-repeat gene family, D6Mm3e; a lysyl oxidase-related gene, Lor2; the evolutionary conserved gene, Aup1; and a novel gene, Gene7. Six genes were eliminated as candidates for mnd2 based on normal mRNA size, abundance, and coding sequence. The remaining three genes--semaM, Lor2, and Gene7--are under continued investigation. The gene order in the mnd2 region between the flanking markers D6Mit164 and D6Mit128 is: centromere - semaphorin M - Gene7 - p62$\rm\sp{Dok}$ - Lor2 - Aup1 - rhotekin - D6Mm3e - dynactin1 - smooth muscle $\gamma$ - actin - telomere. Comparison of different gene identification strategies showed that direct genomic sequencing is the most efficient gene-identification method for this gene-rich chromosomal region. Continued application of the strategy described here is expected to result in identification of the gene that causes the mnd2 phenotype. Elucidation of the molecular defect and its consequences for the pathophysiology of mnd2 mice will broaden our knowledge of molecular mechanisms involved in development and maintenance of the neuromuscular system.Ph.D.Biological SciencesGeneticsMolecular biologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/131232/2/9840562.pd