ANALOGOUS CLEAVAGE OF DNA BY MICROCOCCAL NUCLEASE AND A 1,10- PHENANTHROLINE-CUPROUS COMPLEX. Jessee, G., Gargiulo, G., Razvi, F. and A. Worcel, Nucleic Acids Research 10, Number 19, 5823-5834 (1982).

This study demonstrates the remarkable similarities between the intercalator, 1,10- phenanthroline copper (I), and the micrococcal nuclease in their ability to recognize and to cleave hypersensitive sites in a 5,000 base pair circular DNA fragment containing the histone gene cluster from D. melanogaster.

The figure above shows a comparison between the micrococcal nuclease and the 1,10- phenanthroline copper (I) cleavage patterns, using agarose gel electrophoresis followed by autoradiography. Circularized naked DNA molecules, previously labeled with radioactive phosphorous at a single Bam H1 site, were incubated with either the micrococcal nuclease or 1,10- phenanthroline copper (I), and the reaction followed as a function of time. The resulting fragments were then cleaved with Hind III to give fragments having a common Hind III end, this being 68 base pairs downstream from the labeled Bam site. Slab gel electrophoresis in 1% agarose, followed by autoradiography, was then used to visualize radioactively labeled fragments containing different DNA chain lengths.

Cleavage patterns exhibited by both agents are amazingly similar. Most hypersensitive sites are found at the 5' ends of genes or between adjacent genes.

What is even more remarkable is the observation in subsequent experiments that many of these same sites nucleate melting, with the addition of the single-strand specific DNA binding protein of E. coli in the presence of negative superhelicity in DNA. The location of these small melted DNA regions has been established using S1 nuclease, in combination with electron microscopy (see REFERENCES in main section).