In the sequencing reaction, the reasons for the formation of the templates or primers may lead to the formation of double-peaks.
1) sequencing primers have two binding sites on the template.
2) template is not pure, if it is plasmid or bacterial liquid, the reason is non monoclonal, if it is PCR, the reason is nonspecific band.
3) special structure of template sequence, such as poly structure, hairpin structure, etc.
4) primer degradation, primer impurity, or primer specificity.
