Extraction of RNA from tissues containing high levels of procyanidins that bind RNA |
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Authors: | Chang-Sheng Wang Lila O Vodkin |
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Institution: | (1) Plant and Animal Biotechnology Laboratory, Department of Agronomy, University of Illinois, 61801 Urbana, Illinois, USA;(2) Present address: Department of Agronomy, Taiwan Agricultural Research Institute, 41301 Taichung, Taiwan |
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Abstract: | Commonly used methods for extraction of RNA from plants are not effective for isolation of high quality RNA from the pigmented
seed coats of soybeans that produce procyanidins (tannins) during seed coat development. We demonstrate a significant modification
of the phenol-LiCl method that yields high quality RNA from a black seed coat variety. In this method, seed coat material
was ground in a buffer containing a high concentration of bovine serum albumin (100 mg BSA/50 mg of lyophilized seed coats)
to competitively inhibit proanthocyanidin binding. The presence of hydrated insoluble polyvinylpoly-pyrrolidone (PVPP) was
also necessary to bind proanthocyanidins and remove them from solution. Proteinase K was added to digest the remaining BSA,
and phenol extraction was used to remove both the proteins and small molecular weight complexes formed by BSA and proanthocyanidins.
After LiCl and ethanol precipitations, the RNA quality was examined by UV absorbance spectra, gel electrophoresis, and hybridization.
Using this method, good quality RNA can be extracted from pigmented seed coats of soybean varieties that are homozygous for
the recessivei allele and also contain the dominantT gene that results in production of procyanidins in the seed coat. The method is also effective for tissues from other plant
species that contain abundant polyphenolic compounds. |
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Keywords: | Soybean Glycine max procyanidins flavonoids |
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