I have a PCR product sample which was extracted about 1 year ago. I tried to electrophoresis it and a bright band appeared! Then I diluted it 9 times, 4 times and 2 times into 3 tubes and continued electrophoresis but no band appeared!
Can anyone explain why the second time I did electrophoresis there was no band?
Thank you!
Why did you dilute your PCR product in the first place? Probably after diluting it the concentration is too low to be seen on an agarose gel. However, without seeing the gel picture of the "bright" band this is difficult to judge.
Did you use non-stick plastic tubes? . In your oginal tube it is likely that all the free binding sites were saturated and there was sufficient in solution to visualise on a gel. Diluting into new tubes can sometimes result in the loss of samples sticking to the tubes, particularly if the dilutions are large. I am a bit suprised that with your dilutions you lost everything though.
I diluted due to determine the concentration of it and apply for sequencing. I also used non-stick tubes as Ian Teo confused. And I agree with you, Sabine, a bright band that I got is an obstacle for me to explain.
To overcome the obstacle to explain what a bright band is for you, just upload the gel image.
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