Description
Bbs I is a Type IIS restriction endonuclease encoded by the BbsI gene of Brevibacillus laterosporus. FuniCutTM Series Endonucleases are fast restriction endonucleases that can accurately complete DNA cleavage within 5 to 15 minutes using genetic engineering recombination technology. They are suitable for rapid cleavage of plasmid DNA, PCR products, genomic DNA, etc. FuniCutTM rapid endonucleases share a common enzyme cleavage buffer, simplifying the enzyme reaction system. In addition, they have good enzyme activity redundancy and can easily handle the enzymatic cleavage of excess substrates or difficult templates.
Specifications
|
Cat.No. |
14227ES76/14227ES90/14227ES94/14227ES95 |
|
Size |
500 U/5000 U/20,000U/200,000U |
|
Restriction Enzyme Cut Site |
5'-GAAGAC(N)₂↓-3' |
|
Recommend reaction conditions |
1×FuniCutTM Buffer, Incubate at 37°C |
|
Enzyme activity |
20 U/μL |
|
Inactivation condition |
Incubate at 80℃ for 20 min |
|
Isoschizomer |
BstV2I, BpiI |
Components
|
Components No. |
Name |
14227ES76 (500 U) |
14227ES94 (20KU) |
|
14227-A |
FuniCutTM Bbs I |
25 μL |
1 mL |
|
14227-B |
10×FuniCutTM Buffer |
1 mL |
3 x 1 mL |
|
14227-C |
10×FuniCutTM Color Buffer* |
1 mL |
- |
【Note】: *10×FuniCutTM Color Buffer contains red and yellow tracking dyes which allow the product to be used directly for gel electrophoresis. The red dye in FuniCutTM Color Buffer has a migration rate similar to a 2500 bp double-stranded DNA fragment in 1% agarose gel; the yellow dye has a migration rate similar to a 10 bp double-stranded DNA fragment in 1% agarose gel.
Storage
This product should be stored at -25~-15℃ for 2 years.
Notes
1. Star activity was not observed after 3 h incubation, but delayed enzyme digestion may result in star activity.
2. This product is for research use only.
3. Please operate with lab coats and disposable gloves，for your safety.
Instructions
1. DNA rapid enzyme digestion protocol
1) Prepare the system reaction solution using the following recommended sample addition sequence (operate on ice)
|
Components |
Plasmid DNA |
PCR products |
gDNA |
|
ddH2O |
15 μL |
16 μL |
30 μL |
|
10×FuniCutTM Buffer or 10×FuniCutTM Color Buffer |
2 μL |
3 μL* |
5 μL |
|
Substrate DNA |
2 μL (~1 μg) |
10 μL (~0.2 μg) |
10 μL (5 μg) |
|
FuniCutTM Bbs I |
1 μL |
1 μL |
5 μL |
|
Total |
20 μL |
30 μL |
50 μL |
【Note】：*This system refers to purified PCR products. Unpurified PCR products have a certain ionic strength and the amount of 10×FuniCutTM Buffer added can be reduced accordingly to 2 μL. If cloning and other experiments are performed in the next step, the PCR product must be purified before enzyme digestion.
2) Mix by gentle pipetting or swirling (do not vortex), then centrifuge briefly to collect the droplets.
3) Incubate at 37°C for 15 min (plasmid), 15-30 min (PCR product) or 30-60 min (genomic DNA).
4) Incubate at 80°C for 20 min to inactivate the enzyme and stop the reaction (optional).
5) If FuniCutTM Colour Buffer is used for the digestion reaction, the product can be loaded directly onto the electrophoresis.
2. Double or multiple enzyme digestion protocol
1) The dosage of each endonuclease is 1 µL and the reaction system should be expanded accordingly as needed.
2) The total volume of all endonucleases should not exceed 1/10 of the total reaction system.
3) If the optimal reaction temperatures of the selected endonucleases are different, use the enzyme with the lowest optimal temperature for enzyme digestion, then add the enzyme with the highest optimal temperature and incubate at a higher temperature.
3. Reaction system
|
Components |
Volume（20 μL） |
Volume（20 μL） |
Volume（50 μL）* |
|
DNA |
1 μg |
2 μg |
5 μg |
|
10×FuniCutTM Buffer or 10×FuniCutTM Color Buffer |
2 μL |
2 μL |
5 μL |
|
FuniCutTM Bbs I |
1 μL |
2 μL |
5 μL |
|
Total |
20 μL |
20 μL |
50 μL |
【Note】:*If the total reaction volume is greater than 20 μL, use a water, metal or sand bath and increase the incubation time.
4. Number of recognition sites in different DNA
|
λDNA |
ΦX174 |
pBR322 |
pUC57 |
pUC18/19 |
SV40 |
M13mp18/19 |
Adeno2 |
|
24 |
3 |
3 |
0 |
0 |
3 |
0 |
27 |
5. Effects of methylation modification
|
Dam |
Dcm |
CpG |
EcoKI |
EcoBI |
|
No influence |
No influence |
No influence |
No influence |
No influence |
6. Activity in different reaction buffers*
|
Reaction buffer |
FuniCutTM Buffer |
Thermo Scientific FastDigest Buffer |
NEB CutSmartTM Buffer |
Takara QuickCut™ Buffer |
|
Activity |
100% |
≤10% |
100% |
25% |
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The product is for research purposes only and is not intended for therapeutic or diagnostic use in humans or animals. Products and content are protected by patents, trademarks, and copyrights owned by Yeasen Biotechnology. Trademark symbols indicate the country of origin, not necessarily registration in all regions.
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