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PCR - method (Jun/12/2002 )

Is their any way to open double stranded DNA at lower temperature like 50 to 55 degree?? Any suugestions will be appreciated.


Yes, you can do this with 0.1N NaOH but you won't be able to use the denatured DNA in PCR because it will be too alkaline.


Hi there,
You can add 50%  v/v formamide into your sample. For example, if you have 50 ul DNA solution, you should add 50 ul formamide into that tube, the Tm will be only 50 degC. (This method is used in Southern Blotting). As for PCR, uhm....m, I don't know, why don't you try  with other primers? If the primers are long (>25 bp), you should cut some nucleotides at 3' or 5' terminal which is G-C rich to decrease the Tm.

Best results,

Thai Ke Quan

(Edited by quantk at 7:14 pm on June 13, 2001)