Hi everybody, I'm making a RNA extraction with Trizol from mouse intestinal tissues. In a first step I should break the tissue, I'm using a minidrill with a metal wick for this, the problem is that the trizol changes its color from pink to brown. My question is, this change may affect the collection of cDNA?
If the change in color is due to the tissue then no, this will likely not affect the yield of RNA. However, if the trizol reagent, which is pretty reactive/corrosive, is reacting with the metal wick of the minidrill then this may affect your RNA quality. One method I have used successfully in the past for isolation of RNA from tissues is to freeze the tissue in liquid nitrogen and grind it to a powder in a mortar and pestle. I then add this powder to the trizol in a dounce homogenizer and do a few passes with the homogenizer pestle. This has worked well for me in the past.
If the change in color is due to the tissue then no, this will likely not affect the yield of RNA. However, if the trizol reagent, which is pretty reactive/corrosive, is reacting with the metal wick of the minidrill then this may affect your RNA quality. One method I have used successfully in the past for isolation of RNA from tissues is to freeze the tissue in liquid nitrogen and grind it to a powder in a mortar and pestle. I then add this powder to the trizol in a dounce homogenizer and do a few passes with the homogenizer pestle. This has worked well for me in the past.
Hi, as Zachery said can be caused by tissue itself, for example if you did not wash the tissue in PBS and it remain some blood, or if you have fat in the tissue. After you add the trizol and homogenated the tissue you can perform a centrifuge before the chloroform, recover the super and add the chloroform. I added this step when extract RNA from mouse heart, I had dark coloured samples dued to some blood left.
I fully agree with Zachery. The advatage of tissue homogenisation by grinding to a powder in a mortar and pestle in liquid nitrogen is that the RNA will not become degraded. In contrast, when using a minidrill with a metal wick there is always a danger that some degradation may occur because of insuffcient penetration of Trisol into the tissue. The brown colour of a Trisol reagent may be connected with some local overheating leading to chemical changes of phenol.
Hi, I do Trizol extractions from plant tissue, and get good yields and qualities. However, I do get a brown coloring, which is normal for Trizol. And it may vary depending upon the sample type plant vs animal, and may also vary with different individual tissue type.. even though I am not using any metal for grinding my samples nor do I have any blood involved in the sample.
I had the same outcome with the liver tissue. It was not a problem. The total RNA turned out to be great template for the RACE-PCR.The brown colour is most probably due to pigments carried over from blood during the RNA extraction procedure. But, if it's really bothering, you can try c0-purify the RNA sample through column based RNAeasy kit.
due to brownish or reddish color of the RNA pellet, I usually get bad 260/280 ratio, although concentration is pretty good. Don't know how to solve this problem
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