Dear all, I have used ATMAB protocol for pines DNA extraction. But, after incubation in a heat bath, the solution started black in colour. I applied washed with chloroform for carotenoids but not able to remove the colour. I finished the protocol and made a run on an agarose gel but I don't get an DNA band.
Kindly suggest me some option to tackle the above said problem.
Regards...
Ramiro
Hi Ramiro! I've performed DNA extractions of pine root tips colonized by ectomycorrhizal fungi (looking for fungal ITS). I remember getting a brown color in some of the samples after smashing them in CTAB buffer solution and incubation.. but they amplified well, my guess is that it might be not related with the amplification success.. However, if you have enough sample material maybe adding an extra chloroform wash might work..
Dear Ramiro,
I have the same opinion as Nahuel. Are you sure that the black colour infusing in the solution causing no DNA bands in your AGE?
Have you tried quantifying your DNA yield with a spectrophotometer or Qbit or nano drop? Or tried PCR amplifying a conserved gene like actin? I've never bothered with running out genomic DNA on a agarose gel. You really can't know if the concentration is high enough to use unless you test it empirically.
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