By Ian Marshall, Susan Al-Kazwini, Jenny J. Holman, Peter M. Broad, Mark R. Edbrooke (auth.), G. Fink, A. J. Harmar, Kenneth W. McKerns (eds.)
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Small GTPases play a key function in lots of features of up to date phone biology: keep watch over of telephone development and differentiation; rules of phone adhesion and cellphone circulation; the association of the actin cytoskeleton; and the legislation of intracellular vesicular shipping. This quantity plus its significant other Volumes 255 and 256 conceal all biochemical and organic assays at present in use for studying the position of small GTPases in those features of phone biology on the molecular point.
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As described by M. , 1984), cDNA was prepared after oligo dT priming of polyA enriched RNA with AMV reverse transcriptase in the presence of 100pg/ml Actinomycin D to reduce synthesis of double stranded cDNA. In these studies cDNA was synthesized from both polyA+ RNA of rabbit hippocampus. Removal of relatively abundant sequences common to other brain regions was accomplished by hybridization to a twenty fold excess of mRNA of either cerebellum or cerebral cortex. Following liquid hybridization, remaining single-stranded cDNA was isolated by hydroxyapatite chromatography.
76:449-464. Fonnum, F. , 1978 The effect of intrahippocampal kainic acid injections and surgical lesions on neurotransmitters in hippocampus and septum, J. Neurochem. 31:1173-1181. Gee, C. -L. , Roberts, J. , Thompson, R. and Watson, S. , 1983 Identification of proopiomelanocortin neurones in rat hypothalamus by in situ cDNA-mRNA hybridization, Nature 306: 374-376. Gergen, J. , Stern, R. H. and Wensink, P. , 1979 Filter replicas and permanent collections of recombinant DNA plasmids, Nucleic Acids Res.
Third, this information allows the generation of antibodies with defined sequence specificity for immunohistochemistry to localize the protein produce. intification of cells expressing the mRNA which mayor may not be the same as the protein. Finally, the cloned DNA also provides a hybridization probe for mRNA blots, genomic DNA blots and the isolation of the gene and homologous sequences from a recombinant genomic DNA library. With the cDNA clones presented here and others now being generated in our laboratory and those of others, it is reasonable that the identification of novel and intrinsically interesting proteins will be identified that are germane to the function of the mammalian brain.