Progress in Molecular and Subcellular Biology 3 by A. S. Braverman, D. J. Brenner, B. P. Doctor, A. B.

By A. S. Braverman, D. J. Brenner, B. P. Doctor, A. B. Edmundson, K. R. Ely, M. J. Fournier, F. E. Hahn, A. Kaji, C. A. Paoletti, G. Riou, M. Schiffer, M. K. Wood (auth.), F. E. Hahn (eds.)

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Mean mol. Spec. 4 (1) 16s RNA genes Total Recovery Spec. act. 2 23s RNA genes Total Recovery Spec. " activity (%) (unitS)b b a Nucleotide equivalent. 0. o Spec. , specific activity, m{Lmoles of RNA hybridized per {Lmole of DNA (nucleotide equivalent). • Methylated albumin kieselguhr. 0) DNA {Lg({Lmole)a Purification step Table 5. Purification of the ribosomal RNA genes. (From COLLI and OISHI, 1970) ..... j>. '" 0 :;3 0 4l ...... 0 :;3 r. '"0 ..... ~ 0 42 MAURILLE J. , DON J. BRENNER, and B. P. DOCTOR increased somewhat at the expense of purification.

32P-Iabeled H-strand DNA was then hybridized with unlabeled rRNA and chromatographed on deoxycholate-treated benzoylated DEAE cellulose using a linear NaCl-acetone gradient. Fig. 18 shows the elution profile obtained. 65 M NaCl. In Fig. 18, the bulk of the label elutes as a single broad band mainly in the single stranded DNA region. There is, however, a small shoulder of activity eluting before the main peak (fractions 20 to 35). When fractions were pooled and tested for the ability to rehybridize with labeled rRNA (following removal of the hybridized non-labeled RNA), it was learned that these early fractions were enriched for rDNA.

43 4. 5. 938 Corresponding experiment without any rRNA added 32P_DNA Percent 32p input Cot adsorbed 1. 000125 2. 3. 004 Sununary of results obtained in the modified isolation procedure of rRNA cistrons. Same 32p labeled N. (rassa DNA (190,000 cpm/(Lg DNA) is used in both experiments, (a) control experiment without any rRNA and (b) for making DNA: RNA hybrids. Percent hybridization is measured by calculating the fraction of original input 32p_ N. (rassa DNA adsorbing to hydroxyapatite. , 1972). 01 M phosphate buffer (PB).

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