Altenative way to extract DNA - (Aug/14/2005 )

I tried using DNAzol reagent from Invitrogen to extract DNA from bacteria, but the end product is not as good as those xtraction using kit (such as QIAamp DNA Mini Kit). The concentration is much lower (although I use the same amount of starting material) and PCR is not very good (wondering could it due to the residual ethanol in the DNA samples?). As I'm extracting DNA from a wide range of bacteria, so I would like reduce the cost as much as possible. Any alternative kits of reagents to extract DNA in a fast and yet effective way?

Thanks

Without knowing the species, in general genomic purifications are can be finniky. The pretreatment of cells to lyse can be the key. Lysis with detergents may not be enough. For example, with Staph you have to treat with lysostaphin, then lyse with the given buffer, then purify. With Bacillus, it's not as hardy, but purification and yield is not as efficient as E. coli. Other gram positives require mechanical lysis using beads, shredding agents or homogenizers.

Peruse the literature for methods for your specific species and find better ways to disrupt the memberane and/or cell walls. Then you may be able to get away with using the same kit.

I've heard, but never tried, that it is sufficient to just heat Gram negative strains and use the lysate directly in the PCR. I think there are some thread here in this forum where someone just put a bacterial colony in the PCR tube (with all reagents).

If you are just trying to amplify some portion of the genome, you should be able to drop a colony in your pcr mix. The initial 94-95C steps should lyse the cells. If you need total genomic DNA, try lysozyme for cell lysis with the DNAzol. It's pretty cheap.