Services - DNA Sequencing Electropherogram
Figure 11
Electropherogram that has "the spread".

Bands that gradually spread out and therefore become irresolvable characterize the sequence that has “the spread”. This problem is characterized by delayed migration of the extension products, i.e. the wider the first peak the longer the delay. The first peak that is too broad (irresolvable) can be from base 1 to as late as base 400. The problem is caused by a small, ionic contaminant in the DNA preparation. Most commonly the contaminant is associated with plasmids that were purified with a solid phase extraction kit, e.g. Qiaprep or Wizard. This is a common problem that is not well understood. To solve or minimize the problem the facility will add EDTA to the sequencing reaction, repeat the cleanup procedure, and then analyze the reaction again. When this does not work we repeat the reaction with different reaction conditions to minimize the amount of contaminant and maximize the amount of extension products. At the moment we know of no confirmed methods the customer can do to minimize the contaminant in their plasmid preparation. Since there is little the customer can do to address this problem, then the reactions with this problem are automatically rerun without the need for customers to request that the reaction be repeated.



 Fig. 11a, Electropherogram that has "the spread".


Fig. 11b, Electropherogram that has "the spread".