does glycogen inhibit enzyme digestion or T4 ligation? - (Feb/08/2006 )
thank you very much to al l of you who replied my post
-rose9999_98-
QUOTE (tfitzwater @ Feb 8 2006, 09:50 AM)
Glycogen will not inhibit ligation like yeast tRNA. Does not interfere with A260/280 readings. Glycogen may compete with proteins in nucleic acid:protein interactions. (Aruffo, A. and B. Seed, 1987 Proc. Natl. Acad. Sci. USA 84:8573-8577 and Gaillard, C. and F. Strauss, 1990 Nucl. Acid Res. 18:378.) Glycogen is reported to inhibit the reverse transcription of large templates in a concentration dependent manner. (Baugh, L.R., A.A. Hill, E.L. Brown and C.F. Hunter 2001. Quantitative analysis of mRNA amplification by in vitro transcription. Nucleic Acids Res. 29:E29) Ambion does not recommend the use of glycogen for samples that will be used in a microarray.
Excessive concentrations of tRNA used as a carrier in ethanol precipitation will inhibit ligation reactions. Slight inhibition of colonies per µg DNA at 500 µg/ml tRNA and significant inhibition at 2500 µg/ml. Glycogen shows no inhibition at 1000 or 5000 µg/ml (my data 1986 p 3209).
Excessive concentrations of tRNA used as a carrier in ethanol precipitation will inhibit ligation reactions. Slight inhibition of colonies per µg DNA at 500 µg/ml tRNA and significant inhibition at 2500 µg/ml. Glycogen shows no inhibition at 1000 or 5000 µg/ml (my data 1986 p 3209).
So are there methods to remove glycogen from samples? I've been doing a series of small reactions that require precipitation of small amounts of nucleic acids, and therefore used glycogen... I'm now noticing that when I run my samples on a gel, they're not running normally (which people say could be due to the polysaccharides). I'm wondering if there's a way to rescue my samples by washing away the glycogen.
Thanks in advance.
-olivertam-