RNA Quantitation - Quantitation in presence of Glycogen (Dec/13/2005 )
i need to isolate RNA from very small embryonic tissues, and the quantity use to be so less that i require to add yeast tRNA or GLYCOGEN as a carrier during precipitation step.
While spectrophotometric quantitation these glycogen precipitated RNA samples show an increased O.D. at 260 even if the same dilution is used at different intervals.
As the amount of tissue that i take would give a total yeild of RNA nearly 1ug. But according to the spectrophotometric reading the concentration of RNA comes in a very higher range.
Is the problem associated with the use of Glycogen during the precipitation steps or something else.
Any small suggestion regarding my problem would be a great help..................
don't use tRNA as it willinterfers with your measurements and as you have also glycogen. of cours if you add same tRNA quantity to tubes (and a known one) you can adjusts your measures, but it's better to avoid tRNA in RNA purif.
i ad 1µl of glycogen (100mg/ml) to a 1ml extract.
2 vwashes of ethoh 70% removes quite all glycogen.
I add no pb to quantify with that.