Question

A dideoxy sequencing electropherogram displays bright (high, wide) peaks of fluorescence, obliterating some of the sequencing peaks. What is the most likely cause of this observation? How might it be corrected?

Answer 1

The most likely cause for bright peaks of fluorescence in a dideoxy sequencing electropherogram is the presence of too much dye-labeled ddNTPs. This can be corrected by reducing their concentration in the sequencing reaction.

Step-by-step explanation:

The most likely cause for bright (high, wide) peaks of fluorescence in a dideoxy sequencing electropherogram that obliterates some of the sequencing peaks is the presence of too much dye-labeled dideoxynucleotides. When there is an excess of these labeled ddNTPs, they can result in strong fluorescence, appearing as bright peaks that overpower the signals of the sequencing peaks. This can be corrected by reducing the concentration of the dye-labeled ddNTPs in the sequencing reaction.