Is it better to do an sonification or use a phenol extraction? Which will give me the best results
sonification may be a better way, especially you need to extract active protein.
For protocol, we just keep on sonification, and check whether the mycelium had been smashed under microscope
By the way，if you just need protein for western-blot or other experiment which don`t need protein in active condition.grinding mycelium in liquid nitrogen is a good way,just like RNA-extracting method, and then use water of other buffer to dissolve protein.
I also have some insoluble plant polymer in my media will sonification work ?
Cell lysis is the first step in cell fractionation, organelle isolation and protein extraction and purification. Cell lysis is largely used to obtain the best possible yield and purity for different species of organisms, sample types (cells or tissue) and target molecule or subcellular structure. Historically, physical lysis is the method of choice for cell disruption and extraction of cellular contents; however, it often requires expensive, cumbersome equipment and involves protocols that can be difficult to repeat due to variability in the apparatus (such as loose-fitting compared with tight-fitting homogenization pestles). Also, traditional physical disruption methods are not conducive for high throughput and smaller volumes typical of modern laboratory research.
In recent years, detergent-based lysis methods have become the norm. Through empirical testing by trial and error, different detergent-based solutions composed of particular types and concentrations of detergents, buffers, salts and reducing agents have been developed to provide the best possible results for particular species and types of cells. Detergents have both lysing and solubilizing effects.By careful optimization of physical disruption techniques, detergent-buffer solutions and density gradient methods, procedures have been developed to enable separation of subcellular structures or classes of compounds. For example, with the appropriate detergents, hydrophobic membrane proteins can be solubilized and separated from hydrophilic proteins. A combination of tools and steps enable intact nuclei, mitochondria and other organelles to be isolated for study or protein solubilization. In post lysis condition many critical stages are faced. Cell lysis disturbs the carefully controlled cellular environment, allowing endogenous proteases and phosphatases to become unregulated. As a result extracted proteins become degraded or artifactually modified by the activities of these molecules. Therefore, to prevent these effects and obtain the best possible protein yield in cell lysis, protease and phosphatase inhibitors are added to the lysis reagents. Numerous compounds have been identified and used to inactivate or block the activities of proteases and phosphatases by reversibly or irreversibly binding to them. Lastly to obatin a particular protein, special attention must be given to the effects of the lysis reagents on the stability and function of the protein(s) of interest. Certain detergents will inactivate the function of particular enzymes, and long-term stability of extracted/purified proteins often requires that they be removed from the initial lysis reagents and/or stabilized by addition of particular compounds.
My samples also had some insoluble plant polymer like soya powder in my media and sonification works well, and I removed them by centrifugation after sonification
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