Austin its Gram -ve bacteria...and i want to do molecular analysis on it..
How much DNA do you need? And what kind of "molecular analysis"?
If it is PCR, then all you need to do is touch the colony or culture with a pipette tip and then place it in a PCR mix by pipetting up and down.
Otherwise you need to be more specific about what you are going to do with it, and how much you need.
I have good experience with the DNeasy kit from Qiagen. I used it a couple of times to extract genomic DNA from Gram-positive bacteria to use as a template for PCR. For efficient lysis, I recommend incubating Gram-postive bacteria first in Tris buffer + 10 mg/ml lysozyme at 55 C before proceeding with the lysis step as decribed in the kit.
Now a days no. of companies coming with the good bacterial DNA isolation kit. select any one of them and do your work. Might be nano gram of DNA as a starting templete is enough for the molecular analysis using PCR. If you you want to more precise on costing part, just google it and you can find out various protocol.
Dear Danish,
The following link "Simplification of a Locus-Specific DNA Typing Method (Vir Typing) for Streptococcus pyogenes" worked well for me for PCR amplification of both Gram +ve and Gram -ve isolates.
http://jcm.asm.org/content/36/5/1428
Thankyou Austin ...dear Saanjay singh ji i agreed that there are lots of kits available in the market ...but what i want is to standarize a lysis buffer based method....which can be easier and short method...i dont want to used kit based methods ....Santosh ji iam not getting you sry...link is not woorking..but thanx to all..
Okay.
Then I would look up protocols using alkaline lysis or that use lysozyme + detergent (protocols are in Sambrook and Maniatis). The difference is that you just do the lysis to completion to get the genomic DNA. These methods are particularly useful if you want to harvest intact bacterial genomes (by encasing them in agarose plugs.
Good luck!
Kindly find the attached file. Pg 1428, 1st Column last para. May be useful to you.
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