Isolation and solubilization of proteins after TRIzol extraction of RNA and DNA from patient material following prolonged storage.
A systems approach is being applied in many areas of the biological sciences, particularly in cancer research. The coordinated, simultaneous extraction of DNA, RNA, and proteins from a single sample is crucial for accurate correlations between genomic aberrations and their consequences on the transcriptome and proteome. We present an approach to extract and completely solubilize up to 98% of the total protein recovered from archived samples following TRIzoL isolation of RNA and DNA. We also demonstrate using polyacrylamide gel electrophoresis (PAGE) and Western blot analysis that the proteins, representing both a wide molecular weight range and some posttranslational modifications, such as protein phosphorylation, remain stable in phenol-ethanol for up to 3 years at -20 degrees C.
Hummon
Amanda B
AB
National Cancer Institute, Center for Cancer Research, National Institutes of Health, Bethesda, MD 20892, USA. hummona@mail.nih.gov
Lim
Sharlene R
SR
Difilippantonio
Michael J
MJ
Ried
Thomas
T
D001709:Biotechnology / Q000379:methods*; D004247:DNA / Q000302:isolation & purification; D006146:Guanidines / Q000737:chemistry; D045325:HCT116 Cells; D006801:Humans; D010636:Phenols / Q000737:chemistry; D011506:Proteins / Q000302:isolation & purification*; D012313:RNA / Q000302:isolation & purification; D013048:Specimen Handling
D006146:Guanidines; D010636:Phenols; D011506:Proteins; C411644:trizol; D012313:RNA; D004247:DNA