DNA homology studies in Streptomyces using S1 nuclease

The optimal reaction conditions for the determination of DNA-homology in Streptomyces species were established in the presence of formamide using S1 nuclease. The melting temperature of Streptomyces DNA was 90 degrees C in 0.42 M NaCl containing 20% formamide in which the denaturation was completed by boiling for 5 minutes. In the S1 reaction mixture consisting of 5 U of S1 nuclease, 0.168 M NaCl, 1 mM ZnSO4 and 8% formamide at pH 4.8, single-stranded DNA was hydrolyzed by more than 98%, while the hydrolysis of double-stranded DNA was less than 3%. From the analysis of homoduplex formation, the C0t1/2 was found at 20 hours, when a mixture of unlabeled DNA and index DNA was used at a ratio of 500:1.