S1 nuclease removes 5' and 3' overhanging single-stranded DNA and
hairpin loops. These may occur after restriction enzyme digestions and in cDNA production. Be careful with S1 nuclease. The acidic conditions can depurinate your DNA, and the enzyme may nick double-stranded DNA.
S1 nuclease is compatible with the following buffers:
5X Buffer (source?)
1X BRL Buffer
1x Promega Buffer
1 M NaCl
50 mM NaCl
280 mM NaCl
0.25 M sodium acetate (pH 4.6)
30 mM Na acetate (pH 4.6)
50 mM Na acetate
pH 4.6
pH 4.6
pH 4.5
5 mM ZnSO4
1 mM Zn acetate
4.5 mM Zn sulfate
2.5% glycerol
5% glycerol
Incubate the DNA (about 500 ng/unit of enzyme) at 37°C for 30 minutes. Bring the solution to 20 mM EDTA to stop the reactions. Before phenol extracti
on, adjust the pH of the solution by adding tris to 50 mM.
BRL Tech Line (9/90) reported that mung bean nuclease may be better for blunt-ending a DNA fragment for subcloning. The activity of each is substrate dependent so exact units/µg of DNA can't be given. They suggest 10 U of enzyme for 100 ng of DNA at 15°C for 15-20 minutes in a 50 µl volume. The cooler temperature helps keep the ends annealed.
&
nbsp; Send comments and updates to
Dr. Bart Frank,
Arthritis and Immunology Program, OMRF
Copyright 1993, 1996, and 19
97 by Mark Barton Frank, Ph.D.
Proper citation for data acquired from this document is: "Frank, M. B. S1 Nuclease Digestions. In: Frank, M. B. ed. Molecular Biology Protocols. (http://omrf.ouhsc.edu/~frank/s1.html). 1997. Oklahoma City. Revision Date: October 2, 1997."