RNA isolation - Rna isolation (Jun/08/2004 )
I am trying to isolate RNA from guinea pig liver following the procedure from the paper by Chomczynski and Sacchi.
I do not get RNA on my (formaldehyde) gels and only a very small reading on the photospectrometer despite the positive control working. I have tried the procedure several times.
I just started working, please excuse if this question is too silly, but I really do not know what to do.
The following answers are based upon the beginning/intermediate level:
1. Use Trizol reagents, a modified Chomczynski and Sacchi's method. Follow the kit's manual instruction.
2. One of the centerpieces to get RNA of high quality is to homogenize the tissue efficiently. I prefer to using POLYTRON with proper blade.
3. To judge the quality of RNA, try to run denaturing RNA gel, instead of DNA gel. Also, to put the Ethadium Bromide in the sample/buffer mixture is much better than staining after runing.
To get the RNA from liver should not be a problem.
I got some RNA.
I run RNA on a formaldehyde gel, but other people in the lab told me that the formaldehyde is not necessary, but I can't find anything about denaturating RNA-gels without formaldehyde.