Hi and Good day..
I just want to ask...i deal with staphylococcus species..so, i want to apply boiling method to extract the plasmid because previously i saw some researcher can get plasmid from staphylococcus species or gram positive bacteria..anyone have experience about this?...thank you
Extraction of DNA is not as easy compared to E. coli and other Gram-negatives in most staphylococcal species. To increase lysability of staphylococci we use cell wall digestion with lysostaphin. In our lab we use afterwards a cell disruptung machine (FastPrep) but heat might be suitable in staphylococci with predigested cell wall.
As lysostaphin digest glycin-glycin bridges in the cell wall it is highly active in S. aureus having glycin pentapeptide bridges. In many coagulase negative staphylococci there are several different residues in the cell wall bridge and supplementation of growth media with glycin results in a higher incorporation of glycine and therby a higher susceptibility of the strains against lysostaphin.
The bead extraction method has been a working technique in our lab for many members of Gram-positive bacteria, namely Listeria, Leuconosdoc, enterococcus, etc. It is quick and economic. Please find the method online at
http://www.ncbi.nlm.nih.gov/pubmed/15935495
or
http://www.sciencedirect.com/science/article/pii/S0167701205001077
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