Specific limited cleavage of bihelical deoxyribonucleic acid by wheat seedling nuclease.

Gene-sized pieces produced by digestion of linear duplex DNA with mung bean nuclease

“…In addition to these sites, there are sites in linear duplex DNA that are hydrolyzed more slowly. Under proper conditions, the single-strand specific nuclease from wheat seedlings hydrolyzes X DNA to limit products (Kroeker and Fairley, 1975), the distribution of which shows that cleavages are made in the left half of the molecule as well as in the right half which contains the A + T rich blocks.…”

Mung bean nuclease I. 7. Terminally directed hydrolysis of native DNA

“…Also, at the times indicated, 0.01 ml was removed and diluted to 0.1 ml to attain a final Tris-HCl concentration of 0.02 M, pH 8.1. Neutral sucrose gradient sedimentation was performed with these samples and molecular weight were calculated as previously described (Kroeker and Fairley. 1975).…”

“…The limited endonucleolytic activity of mung bean nuclease on native T7 DNA suggests the presence of a small number of specific, susceptible sites on the T7 genome to the action of the enzyme. This observation has been made for a variety of DNA that were cleaved to a limited extent by another "single-strand specific" enzyme, wheat seedling nuclease (Kroeker and Fairley, 1975). The physical and chemical nature of these sites, as well as the mechanism in which they are recognized and cleaved by mung bean nuclease, are currently being investigated.…”

“…In addition, it has been shown that Si nuclease can hydrolyze the phosphodiester bonds involved in small mismatched regions (possibly at the level of a single base pair) produced by mutational alteration in the DNA duplex (Shenk et al, 1975). Kroeker and Fairley (1975) have shown that wheat seedling nuclease is capable of cleaving intact bihelical bacteriophage DNA to a limited extent producing double-stranded polymers, 2-3 X 106 daltons, which are resistant to further action by the enzyme. Mung bean nuclease has been used on naturally occurring native DNA as a structural probe of superhelical DNA (Wang, 1974) and as a structure-function probe of the binding of repressor protein to -phage DNA (Chan and Wells, 1974).…”

“…Gel electrophoresis was performed as described in the preceding paper of this issue (Kowalski et al, 1976). Sedimentation analysis of T7 DNA in neutral and alkaline sucrose gradients was done as previously described (Kroeker and Fairley, 1975).…”