Analysis of repetitive and unique sequences in the mouse genome using fluorescence in situ hybridization and digital imaging microscopy

I have used fluorescence in situ hybridization in combination with digital imaging microscopy (using either a CCD camera or a confocal laser scanner) to explore the organization of DNA sequences in metaphase chromosomes and interphase nuclei of mouse cells. The mouse genome contains both unique sequences and interspersed and tandemly repeated DNA elements. The murine interspersed repeats include the LINE-1 sequence, and two short repeats, the B1 and B2 elements. The distribution of the mouse LINE and SINE sequences on metaphase chromosomes was defined by fluorescence in situ hybridization. The LINE sequences reside in Giemsa (G) bands, while the SINE repeats occupy reverse (R) bands. The L1 sequences in Mus spretus and Mus castaneous mice were also shown to reside in G bands.;Cytogenetic mapping of murine unique sequence clones has been hampered by the inability to easily identify the chromosomes. Hybridization of mouse L1 sequences results in Giemsa-like bands that permit chromosome karyotyping and band assignment of a cohybridized probe. L1 hybridization and DAPI staining were used to map a number of genes previously localized by genetic methods, as well as twenty new clones. A collection of marker clones mapping to each chromosome was assembled to use for identification of chromosomes when the nature of the metaphase preparation makes analysis difficult.;While chromosome specific repeat sequences have been isolated for most human chromosomes, there is only one mouse repeat, other than the repetitive elements on the Y chromosome, (on chromosome X) described in the literature. I identified a second repeat which maps below the centromeric region on chromosome 8. The sequence displays high chromosomal and species specificity. Evidence suggesting that the repeat unit is large and is arranged in an interspersed fashion is discussed.;Another repeat sequence mapping to a similar region on chromosome 14 also was identified. Experiments which parallel the studies on the chromosome 8 repeat have been initiated. A discussion of the similarities of these sequences with the published X repeat is presented.