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The Journal of Cell Biology
The Rockefeller University Press
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766b.tif THE ISOLATION OF ENZYMICALLY ACTIVE NUCLEI FROM THE RAT HEART AND UTERUS C. C. WIDNELL, T. H. HAMILTON, and J. R. TATA. From the National Institute for Medical Research, Mill Hill, London, England. Dr. Widnell's present address is The Rockefeller University, New York. Dr. Hamilton's present address is Department of Zoology, University of Texas, Austin Recent results obtained in many laboratories (see I) have focused attention on the role of the cell nucleus in the action of growth-promoting hor- mones. In order to extend our studies on the action of thyroid hormone, growth hormone, and testosterone on liver RNA polymerase (2, 3) to other tissues, we have modified our method for the isolation of rat liver nuclei (4). We report here a method for the isolation of enzymically active nuclei from the rat heart and uterus. EXPERIMENTAL Isolation of Heart Nuclei All procedures were performed at 0-4 °. Hearts from at least five malc Sprague-Dawlcy rats (150 -4- 20 g body weight) were pooled and weighed in 0.32 sucrose, 3 lnM MgC12 (homogenizing medium, sec 4); the tissue was minced finely with curved scissors and passed, using a stainlcss steel tissue press, twice each through sieves with pore diameters of 1.27 and 0.95 ram. The fragmented tissue was then homoge- nized in 3 volumes of homogenizing medium for 2 min 30 scc using an Ultra Turrax TP 18/2 tissue disintegrator (Janke und Kunkel K.G., Staufcn 1. BR., Germany) run at 60 v (mains, 240 v). The homogenate was filtered through two layers of nylon bolting cloth (110 mesh, John Staniar Ltd., Man- chester, England) and the nylon bolting cloth rinsed with an equal volume of homogenizing medium. 25 ml of homogenate were diluted with 7 ml of water, 15 ml of homogenizing medium were layered under- neath, and a crude nuclear fraction was isolated by centrifugation at 700 g for 10 min. The pellet was re_suspended to a final volume of 13 ml in 2.4M sucrose 1 mM MgCl2 using a ground-glass homo

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