According to enzymatic method for extracting extracellular DNA in biofilm matrix, is suggested to use CTAB solution. Why? I only need extracellular DNA...
Do you have a similar protocol to extract eDNA?
The optimum protocol for extracellular DNA extraction from biofilm can vary depending on the specific characteristics of the biofilm and the intended downstream applications of the extracted DNA. However, here is a general protocol that can be used as a starting point:
Collect the biofilm sample and transfer it to a sterile tube. Add an appropriate volume of phosphate-buffered saline (PBS) to the tube to resuspend the biofilm.
Sonicate the sample using a probe sonicator to break up the biofilm and release the extracellular DNA. Alternatively, vortex the sample vigorously for several minutes.
Centrifuge the sample at a low speed (e.g., 5000 rpm) for 10-15 minutes to pellet the cells and large debris.
Transfer the supernatant containing the extracellular DNA to a fresh sterile tube.
Add a protease (e.g., proteinase K) and an appropriate volume of buffer to the sample to lyse any remaining cells and release the intracellular DNA. Incubate the sample at an appropriate temperature (e.g., 55°C) for an appropriate amount of time (e.g., 1 hour).
Extract the DNA from the sample using a commercial DNA extraction kit or a phenol-chloroform extraction protocol.
Quantify the extracted DNA using a fluorometer or a spectrophotometer.
It's important to note that extracellular DNA can be prone to degradation, so it's important to minimize the time between sample collection and DNA extraction and to store the sample and extracted DNA appropriately. Additionally, it may be necessary to optimize this protocol for the specific characteristics of your biofilm sample and downstream applications.
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