Recommended
buffer |
Reaction conditions |
|
Temperature |
Time |
| FastDigest® buffer |
37°C |
5 min |
Thermal Inactivation
FastDigest® AflII (BspTI) is inactivated by incubation at 65°C for
10 min.
Digestion of Plasmid DNA
1 µl of FastDigest® AflII (BspTI) digests up to 1 µg of plasmid DNA in 5 min, see
Protocol for Fast
Digestion and Analysis of Plasmid DNA with FastDigest® Enzymes
and
Protocol for Fast Vector
Preparation with FastDigest® Enzymes.
Digestion of PCR Products
1 µl of FastDigest® AflII (BspTI) digests ~0.2 µg of PCR product in 5 min,
see
Protocol for Fast Digestion of PCR
Products with FastDigest® Enzymes.
Digestion of Genomic DNA
1 µl of FastDigest® AflII (BspTI) digests 1 µg of genomic DNA in 5 min, or 5 µg of
genomic DNA in 30 min, see
Protocol for Fast Digestion of Genomic DNA with FastDigest® Enzymes.
Ligation
and Recleavage
After overdigestion with 1 µl of FastDigest® AflII (BspTI)
for 1 hour, more than 95% of the digested phiX174 DNA fragments can be ligated in a reaction mixture containing 20-40 u of
T4 DNA Ligase/1 µg of fragments
and 10% PEG. More than 95% of these can be recut.
| Methylation
Effects |
Dam:
Dcm:
CpG:
EcoKI:
EcoBI: |
never overlaps
never overlaps
never overlaps
never overlaps
never overlaps |
- no effect.
- no effect.
- no effect.
- no effect.
- no effect. |
Recommendations for Double and Triple Digestions with FastDigest® Enzymes
Perform 5 minute double or triple digestions with FastDigest®
AflII (BspTI) and any other FastDigest® enzyme(s) in FastDigest® buffer,
see Protocol for
Double Digestion with FastDigest® Enzymes and
Protocol for Triple Digestion with
FastDigest® Enzymes.
