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Plant and Cell Physiology, 1985, Vol. 26, No. 8 1499-1509
© 1985

Article

Quantitative Isolation of Undegraded Polysomes from Aged Pea Tissue in the Absence of Contaminants and Artefacts

Shunnosuke Abe1 and Eric Davies

School of Biological Sciences, University of Nebraska Lincoln, Nebraska, 68588-0118, U.S.A.

Undegraded polysomes were isolated successfully from aged pea epicotyls by grinding frozen tissue in at least 10 volumes of buffer A (0.2 M Tris-HCl, pH 8.5; 0.2 M sucrose; 60 mM KCl; 30 mM MgCl2), taking care to prevent the tissue from thawing prior to homogenization. Supposedly pure polysomes, derived from the membrane-bound polysome fraction, were apparently contaminated with membranes, and contained polysomes clumped together via nascent chains. Problems with contaminants and artefacts were partially alleviated by the use of polyoxyethylene tridecyl ether as a detergent replacing Triton X-100; further alleviated by the use of large volumes of detergent-containing buffer to resuspend the membrane-bound polysome; and almost completely eliminated by brief treatment of resuspended polysomes with protease K. Optimal conditions for isolating polysomes from aged tissue are given.

1 Present address: Institute of Agricultural Environment Control, College of Agriculture, Ehime University, 3-5-7 Tarumi, Matsuyama 790, Japan.

(Received April 24, 1985; Accepted September 2, 1985)
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