An improved method for the purification of higher plant ribosomal RNA labeled on the methyl groups
✍ Scribed by J.P. Cecchini; R. Miassod
- Publisher
- Elsevier Science
- Year
- 1979
- Tongue
- English
- Weight
- 663 KB
- Volume
- 100
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
None of the methods already reported for elimination of pectins from rRNA extracts allowed the complete removal of methylated polysac~ha~des from methyI-labeled cytoplasmic 17 and 26 S rRNA preparations of sycamore (Acer pseadoplatanas L.) cells. An improved procedure for purifying large amounts of higher plant cytopiasmic rRNA labeled on the methyl groups was investigated. Bulk cellular RNA from sycamore cells incubated for 24 to 36 h with methyl-labeled methionine was extracted at 4°C by the phenol-extraction procedure. Most of the pectic compounds (that accounted for about 30% of the total label of RNA extracts) was selectively precipitated, before the 66% ethanol precipitation of nucleic acid, by bringing the deproteinized aqueous layer to 10% ethanol-O. i5 M sodium acetate. Cytoplasmic rRNA, 17 and 26 S, were isolated by repeated sucrose gradient sedimentations and further chromato~aphed on a methylated albumin kieselgurh (MAK) column. The old-fashioned MAK ~hromatog~phy proved to be very useful for eiimin~on of residual pectins, since these compounds eluted in the void volume of the column. This purification procedure gave in a reproducible way cytoplasmic 17 and 26 S rRNA virtually free of any labeled DNA, mRNA, plastid rRNA, andbe& compounds.