5'...G A G C T↓C...3'3'...C↑T C G A G...5'
Lambda DNA digested with SacI, 0.7% agarose, 2 cleavage sites
Thermo Scientific conventional restriction endonucleases are a large collection of high quality restriction enzymes, optimized to work in one of the buffers of the Five Buffer System. In addition, the universal Tango buffer is provided for convenience in double digestions. All of the enzymes exhibit 100% activity in the recommended buffer and reaction conditions. To ensure consistent performance, Thermo Scientific restriction enzyme reaction buffers contain premixed BSA, which enhances the stability of many enzymes and binds contaminants that may be present in DNA preparations.
SacI is sensitive to cytosine methylation at GAGmCTC, but not GAGCTmC and insensitive to adenine methylation at GmAGCTC. AluI methyltransferase (AGmCT) can be used to block SacI. Supercoiled plasmids may require up to 5-fold more SacI for complete digestion than linear DNA.
SacI is inhibited by common clinical anticoagulants found in some preparation of anticoagulated peripheral blood and bone marrow. Levels of EDTA and ACD (citric acid-sodium citrate-dextrose) in standard sample preparation have been shown to inhibit SacI. Three times the normal concentration for heparin is required to inhibit SacI. (J. E. Coad et al., Inhibition of restriction endonucleases by common clinical anticoagulants. Anal. Biochem. 205, 368-369, 1992).
For methylation sensitivity refer to product specifications.
m5C GAGCTm5C G