Previously added items:
5'...m5C N N G (N)9 ↓...3'3'... G N N C (N)13↓...5'
pBR322 DNA (dcm+) digested with SgeI, 1.4% agarose, 6 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.
DNA methylated by Dcm or CpG methyltransferases will be a substrate for SgeI. Greater than 3-fold overdigestion with SgeI may result in incomplete cleavage. At least two copies of SgeI recognition sequence are required for an efficient cleavage. Amount of the enzyme required for complete digestion of methylated DNA depends on the number of SgeI recognition sites. DNA cleavage products generated by target site cleavage facilitate the nonspecific cleavage by SgeI.
Therefore, optimization of the enzyme amount is recommended for DNA cleavage. pBR322 DNA isolated from E. coli dcm+ strain (#SD0041) can be used as a DNA cleavage efficiency control. SgeI cleaves all six dcm methylated targets on pBR322 DNA.
For methylation sensitivity refer to product specifications.
m5C Gm5C G