We describe the reaction of formamide with 2’-deoxyadenosine and 2’-deoxyguanosine to give imidazole ring opening by nucleophilic addition on the electrophilic C(8)-position of the purine ring. This information allows improvement of the one-lane chemical DNA sequencing procedure based on the base-selective reaction of formamide with DNA. The reactivity with formamide of several 7-deazapurine analogues (7-deaza-2’-deoxyinosine, 7-deaza-2’-deoxyguanosine, and 7-deaza-2’-deoxyadenosine) incorporated into polynucleotides is also described. The wide spectrum of different sensitivities to formamide displayed by these purine analogues provides the single-lane DNA chemical sequencing procedures with the possibility of wide-ranging signal intensity modulation and thus increased specificity.