I am currently working on extracting dna from H. pylori culture isolates by phenol-chloroform-isoamyl alcohol method by using CTAB/NaCl. Can I use the same method of extraction but without using CTAB? If yes, then how?
Yes you can use Glucose,Tris EDTA without adding CTAB in your extraction buffer
Thank u for the suggestion. While using phenol chloroform isoamyl alcohol in the extraction process, my phases do not separate well. I do not get a clear aqueous phase. what must be the reason? can anyone plz suggest...?
hi Anisha Sarma, yes we can isolate DNA without using CTAB, which we usually follow in our lab. our protocol includes first treating with Tris, EDTA, Glucose buffer followed by lysis buffer (lysozyme or SDS) followed by RNase treatment and next step is phenol chloroform isoamyl alcohol treatment, where we can see the clear phases after centrifugaton. after that ice cold isopropanol step, alcohol and final suspension step. we have done many times and we are successful in isolation , its usage further in molecular work also.
Note: I have explained the protocol in brief. the concentrations and pH of buffers, centrifugation speed was mentioned.
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