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Potvrzuji
  1. Úvodní strana
  2. Reagencie
  3. SgeI

SgeI

Kód produktu: ER2211 Kód výrobce: ER2211 Kód dodavatele: {2C4618C6-6877-4D00-9164-3026D62DDE7E} Výrobce: UAB Fermentas
SgeI
na dotaz
250 Units

5'...m5C N N G (N)9 ↓...3'
3'...  G N N C (N)13↓...5'

Zobraz detailní popis

Detailní popis

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.

Highlights

  • Superior quality – stringent quality control and industry leading manufacturing process
  • Convenient color-coded Five Buffer System
  • Includes universal Tango buffer for double-digestions
  • BSA premixed in reaction buffers
  • Wide selection of restriction endonuclease specificities

Applications

  • Molecular cloning
  • Restriction site mapping
  • Genotyping
  • Southern Blot
  • Restriction fragment length polymorphism (RFLP)
  • SNP

Note

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.

Conditions for 100% Activity
  • 1X Buffer SgeI: 10 mM Tris-HCl (pH 8.0 at 37°C), 5 mM MgCl2, 100 mM KCl, 0.02% Triton X-100, and 0.1 mg/mL BSA
  • Incubate at 37°C
Digestion of Agarose-embedded DNA Minimum 3 units of the enzyme are required for complete digestion of 1 µg of agarose-embedded pBR322 DNA isolated from E. coli dcm+ strain DNA in 16 hours at 37°C.
Double Digestion Perform double digestion using DoubleDigest.
Hazardous No
Isoschizomers Search for commercial isoschizomers using REsearch.
Storage Buffer SgeI is supplied in: 10 mM Tris-HCl (pH 7.4 at 25°C), 100 mM NaCl, 1 mM DTT, 1 mM EDTA, and 50% (v/v) glycerol.
Storage Condition -20 C

Reaction conditions

Recommended buffer for 100% activity Optimal temperature Enzyme activity in Thermo Scientific buffers, % Tango buffer for double digestion
B (blue)
1X		 G (green)
1X		 O (orange)
1X		 R (red)
1X		 Tango
(yellow)
1X / 2X
SgeI Buffer (Unique) 37°C 0-20 0-20 0-20 NR NR NR NR

Methylation Effects

  • Dam: never overlaps - no effect.
  • Dcm: always cleaves DNA methylated by Dcm methyltransferase.
  • CpG: cleaves targets overlapping with CpG methylated sequences.
  • EcoKI: never overlaps - no effect.
  • EcoBI: never overlaps - no effect.
Methylation type Sequence Cleavage effect
Dcm (CCWGG) Cm5CWGG Cleaves only methylated DNA
CpG 5'...m5C GNG...3'
3'... Gm5CNC...5'		 Cleaves only methylated DNA
CpG 5'...m5C Gm5C G...3'
3'... Gm5CGm5C...5'		 Cleaves only methylated DNA

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