. Embedment-free electron microscopy shows the nuclear matrix comprised of a network of 11 nm filaments with numerous embedded dense organelles. They will use two new nuclear matrix preparations, high salt and the stabilized pre- fixed matrix. They will determine the true nature of the interchromatin granules using reinless section EM of the nuclear matrix preparations immunostained with antibodies to splicing components (in collaboration with Dr. Phil Sharp laboratory). They will combine these results with in situ hybridization with intron specific probes (in collaboration with Dr.Jeanne Lawrence) to locate which nuclear matrix organelle(s) are the location of splicing. In situ hybridization of hnRNA tracks will image the RNA transport machinery. Hybridization with an intermediate repetitive sequence will identify the location of hnRNA that does not serve as mRNA precursor. They will seek monoclonal antibodies to NM filament proteins to determine their number, molecular sequences and fate at mitosis.
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|Nickerson, J A; Krockmalnic, G; Wan, K M et al. (1997) The nuclear matrix revealed by eluting chromatin from a cross-linked nucleus. Proc Natl Acad Sci U S A 94:4446-50|
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