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Simultaneous imaging and classification of all chromosomal territories in the 3D interphase nucleus using spectral karyotypingAbstractMajor success has been achieved in recent years in understanding the function of the genome. It also became evident that the structure and organization of the genome in the nucleus is important, it changes along the cell cycle and during cancer progression. One of the more recent structures found in mammalian nuclei is the chromosomal territory (CT). The chromatin of every chromosome has a distinct and non-overlapping space in the 3D interphase nucleus. Many research questions have arisen regarding these CTs. In a recent study (Louis, Vermolen et al. 2005), for example, we showed a relationship between breakage-bridge-fusions, telomere aggregates and CT overlaps. In this presentation we will introduce a novel technique to image and classify all CTs simultaneously in the 3D interphase nucleus. The basis of this technique relies on a proven technique known as spectral karyotyping (Garini, Macville et al. 1996). Here the chromosomes are labeled with five dyes using a combinatorial table (enough combinations can be made to label each chromosome uniquely) and imaged using a Sagnac interferometer, providing a spectrum for every pixel of the CCD. The classification is based on a linear decomposition of the colors used to probe the chromosomes. For the 3D SKY we use the same hardware to acquire 3D spectral images. At different focal planes a spectrum is acquired after which the images need to be deconvolved and then classified. We foresee this technique as being an excellent instrument for research on chromosomal territories.
Louis, S. F., B. J. Vermolen, et al. (2005). "c-Myc induces chromosomal rearrangements through telomere
and chromosome remodeling in the interphase nucleus." PNAS 102(27):9613-9618. |