Comments
The high concentration of polyphenols in many plants is a problem for isolating nucleic acids, as these substances will inhibit enzymes in the following procedures. They can be absorbed by the addition of 1 % polyvinylpyrolidone K30 (v/v; final concentration) during the tissue homogenization (ref. 1 page 2.3.4). PVP should be prepared fresh as a 20 % stock each time (2).Positively charged nylon membranes used in immunoblots show a high non-specific background. Therefore, these surfaces have to be blocked, in case of the nylon membrane in a very stringent way. Casein (6 % w/v) in combination with 1 % (v/v) polyvinylpyrrolidone (PVP) can be applied. Both substances are dissolved by stirring for 5 minutes in warm (65°C) TTBS buffer.
Literature
(1) Ausubel, F.A., Brent, R., Kingston, R.E., Moore, D.D., Seidman, J.G., Smith, J.A. & Struhl, K. (eds.) (1995) Current Protocols in Molecular Biology. Page 2.3.4 (Suppl. 27) and Page 10.8.15 (Suppl. 39). Greene Publishing & Wiley-Interscience, New York. (2) John, M.E. (1992) Nucleic Acids Res. 20, 2381. Isolation of RNA and DNA from plants with a high content of polyphenols.
