High-yield skeletal muscle protein recovery from TRIzol after RNA and DNA extraction

Extraction of DNA, RNA and protein from the same sample would allow for direct comparison of genomic, transcriptomic and proteomic information. Commercially available kits exhibit poor protein yield and the TRIzol(R)reagent produces a protein pellet that is extremely difficult to solubilize. In response to these limitations, this study presents an optimized method for the extraction of protein from the organic phase of TRIzol that allows for higher yield recovery of skeletal muscle protein compared with direct homogenization in a common protein lysis buffer. The presented method is inexpensive, simple and fast, requires no additional treatment of the protein pellet for dissolution, and is compatible with downstream western blot applications. Lay abstract Scientists analyze DNA, RNA and protein using separate kits and techniques that do not allow for effective analysis of all three macromolecules from the same sample. Simultaneous extraction kits and techniques are limited by poor protein yield after nucleic acid isolation. We present a fast, effective, inexpensive and high-yield method of recovering protein (including large proteins such as titin) from tissue using the TRIzol reagent after RNA and DNA recovery. Method summary The method of high-yield protein extraction from TRIzol after RNA and DNA isolation involves replacing chloroform with bromochloropropane. Instead of producing a tightly packed protein pellet using isopropanol, the protein in the organic phase is precipitated using ethanol and water. Complete dissolution of the resulting protein pellet is achieved using a sodium dodecyl sulfate-urea buffer that allows solubilization of large protein species.