I have treated my cyanobacterial pellets with lysis Buffer (50mM HEPES + 300mM NaCl, pH 7.5). After adding lysis buffer I have sonicated for 20 mints in the presence of ice. After sonication i have centrifuged the lysate mixture with 8000rpm for 45 mints. After centrifugation the supernatant are still green not blue as usually seen for total proteins. What should be the problem ? Lysis is not done completely ?
I doubt about your sonication time ! You will have to work closely on this method by breaking the cycle in multiple numbers and cooling it down alongside ( lets say 5 sec or 10 sec whatever time would give you maximum cell protein). or You can also use beads and vortexing the cells in lysis buffer.
Just check the following link would help you to get your results precisely:
Article Protein Extraction, Fractionation, and Purification From Cyanobacteria
I wonder if you tried including a detergent as for example, Tritonx100 in your lysis buffer? If you haven't tried this, try it & compare the results, to ensure that the lysis is complete.
Meantime be careful with the method that you follow in sonication since it should be at intervals of 15 seconds on and 30 seconds off for the required period, even though you are doing the process using an ice bath since the prob of the sonicator will heat up to a very high temperature that may lead to losing some molecules including the one you are looking for.
Good luck.
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