PstI
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
Conditions of high pH, low salt, high glycerol, 8% DMSO can cause star activity (Malyguine, E., et al., Gene, 8, 163-177, 1980). Surrounding sequences: the presence of adjacent runs of G-C base pairs confers significant resistance to cleavage (K. Armstrong, W.R. Bauer. NAR 10, 993-1007, 1982). 100% dUTP incorporation at the recognition site reduces PstI cleavage to 25% (T.C. Glenn et al., Biotechniques. 17, 1086-1090, 1994). PstI will not cut AGCTGCAG when methylated by AluI methyltransferase.
For methylation sensitivity refer to product specifications.
| Compatible Ends | Alw21I, BseSI, Mph1103I, PstI, SdaI. |
| Conditions for 100% Activity |
|
| Digestion of Agarose-embedded DNA | Minimum 5 units of the enzyme are required for complete digestion of 1 µg of agarose-embedded lambda DNA in 16 hours. |
| Double Digestion | Perform double digestion using DoubleDigest. |
| Hazardous | No |
| Isoschizomers | Search for commercial isoschizomers using REsearch. |
| Storage Buffer | PstI is supplied in: 10 mM Tris-HCl (pH 7.4 at 25°C), 200 mM NaCl, 1 mM DTT, 0.1 mM EDTA, 0.15% Triton X-100, 0.2 mg/mL BSA, 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 |
||||
| O (Orange) | 37°C | 50-100 | 50-100 | 100 | 100 | 50-100 | 50-100 | 1X or 2X |
Methylation Effects
- Dam: never overlaps - no effect.
- Dcm: never overlaps - no effect.
- CpG: never overlaps - no effect.
- EcoKI: never overlaps - no effect.
- EcoBI: never overlaps - no effect.
Cleavage efficiency close to the termini of PCR fragments
| bp from the recognition site to fragment end | ||||
|---|---|---|---|---|
| 1 | 2 | 3 | 4 | 5 |
| 0-20 | 50-100 | |||
Number of recognition sites in DNA molecules
| Lambda | ΦX174 | M13mp18/19 | pBR322 | puc18/19 | pUC57 |
|---|---|---|---|---|---|
| 28 | 1 | 1 | 1 | 1 | 1 |
| pTZ19R/U | pTZ57R | pBluescriptIIKS(±) | pBluescriptIISK(±) | pACYC177 | pACYC184 |
| 1 | 1 | 1 | 1 | 1 | 0 |
New sites generated by ligation
Newly generated recognition sites resulting from ligation of protruding compatible DNA ends
| Recognition Sequence | Second Enzyme | Enzymes recognizing newly generated recognition sequence |
|---|---|---|
CTGCA^G |
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