Support, restriction enzymes: Fermentas Five Buffer System; Dilution of Restriction Enzymes

Restriction Enzymes

Modifying Enzymes

PCR, qPCR, RT-PCR & dNTPs

Molecular Cloning

Nucleic Acid Purification

Molecular Labeling & Detection

In vitro Transcription

Electrophoresis Products

Nucleotides

Transfection Reagents

Reagents

Reaction Conditions for Restriction Enzymes:

General Protocol for DNA Digestion

Five Buffer System (see below)

Dilution of Restriction Enzymes

Stability During Prolonged Incubation

Inactivation

Considerations for Partial Digestion of DNA

Chart: Reaction Conditions for Restriction Enzymes

Five Buffer System

Our unique Five Buffer System features the optimum reaction conditions for each restriction enzyme. The system consists of B (blue), G (green), O (orange), R (red) and Tango™ (yellow) buffers (for buffer compositions, please see table below). All restriction enzymes are packed in color-coded tubes to indicate the recommended reaction buffer. The 10X recommended buffer and/or the universal 10X Tango™ buffer are supplied with each enzyme. Tango™ buffer has been specifically designed for double digestions. For more information on double digestion, please refer to the section “How do I Perform Double Digestion?” or visit the Fermentas DoubleDigest™ engine.

The Five Buffer System ensures optimal enzyme performance, simplicity and convenience. To ensure dependable and reproducible restriction endonuclease performance, Fermentas buffers contain BSA, which enhances the stability of many restriction endonucleases and binds contaminants that may be present in DNA preparations. Due to the stringent requirements involved in our BSA preparations, Fermentas buffers containing BSA can be freeze-thawed multiple times without BSA precipitation.

Fermentas restriction enzymes exhibit 100% of their certified activity in the recommended buffer. Some restriction enzymes require additives to achieve 100% activity. For example, AjuI, AlfI, BdaI, BplI, BseMII, FaqI, Eco57I, Eco57MI, Hin4I, TsoI require S-adenosylmethionine, which is supplied with the enzyme, while AarI and BveI require oligonucleotide (also supplied with the enzyme), and Esp3I requires DTT (DTT is not stable in solution. A freshly made DTT solution should be added directly to the reaction mixture before digestion).

The following enzymes require unique buffers for optimal digestions: AarI, AjiI, BamHI, BfuI, Bpu10I, BseXI, Bsp143I, Cfr9I, Cfr10I, Eam1105I, Ecl136II, Eco52I, EcoRI, KpnI, PasI, Ppu21I, SacI, ScaI, SdaI, SduI and TaqI. For the compositions of unique buffers, see table below, or see the descriptions of restriction enzymes in the Certificate of Analysis provided with each enzyme.

Recommended storage conditions. All Fermentas buffers with BSA should be stored at -20°C.

Reaction Buffers for Restriction Enzymes

The buffers listed below are available from Fermentas and may be ordered separately from the restriction enzymes.

Fermentas Buffer 1X Buffer Composition Catalog # Quantity
Certificate of Analysis (in pdf, ~20 KB), MSDS

Buffer Set for Restriction Endonucleases B30 1 ml 10X each of B, G, O, R, Tango™ buffers.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer B (blue) 10 mM Tris-HCl (pH 7.5 at 37°C),
10 mM MgCl₂,
0.1 mg/ml BSA BB5 5x1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer G (green) 10 mM Tris-HCl (pH 7.5 at 37°C),
10 mM MgCl₂,
50 mM NaCl,
0.1  mg/ml BSA BG5 5x1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer O (orange) 50 mM Tris-HCl (pH 7.5 at 37°C),
10 mM MgCl₂,
100 mM NaCl,
0.1 mg/ml BSA BO5 5x1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer R (red) 10 mM Tris-HCl (pH 8.5 at 37°C),
10 mM MgCl₂,
100 mM KCl,
0.1 mg/ml BSA BR5 5x1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer Tango™ (yellow) 33 mM Tris-acetate (pH 7.9 at 37°C),
10 mM magnesium acetate,
66 mM potassium acetate,
0.1  mg/ml BSA BY5 5x1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for BamHI, Lsp1109I 10 mM Tris-HCl (pH 8.0 at 37°C),
5 mM MgCl₂,
100 mM KCl,
0.02% Triton X-100,
1 mM 2-mercapthoethanol,
0.1 mg/ml BSA B57 5x1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for BfuI 50 mM Tris-acetate (pH 7.9 at 37°C),
15 mM Mg-acetate,
100 mM K-acetate,
0.1  mg/ml BSA B59 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for AarI, AjiI, Bpu10I, ScaI, PasI 10 mM Bis-Tris Propane-HCl (pH 6.5 at 37°C),
10 mM MgCl₂,
100 mM KCl,
0.1 mg/ml BSA B27 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for BseXI
50 mM Tris-HCl (pH 7.5 at 37°C),
2 mM MgCl₂,
100 mM NaCl,
0.1 mg/ml BSA B31 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for Bsp143I 33 mM Tris-acetate (pH 7.9 at 37°C),
10 mM Mg-acetate,
66 mM K-acetate,
0.02% Triton X-100,
0.1 mg/ml BSA B13 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for Cfr9I 10 mM Tris-HCl (pH 7.2 at 37°C),
5 mM MgCl₂,
200 mM sodium glutamate,
0.1 mg/ml BSA B02 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for Cfr10I 10 mM Tris-HCl (pH 8.0 at 37°C),
5 mM MgCl₂,
100 mM NaCl,
0.02% Triton X-100,
0.1 mg/ml BSA B04 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for Eam1105I 10 mM Tris-HCl (pH 7.5 at 37°C),
5 mM MgCl₂,
100 mM NaCl,
0.1 mg/ml BSA B25 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for Ecl136II, SacI 10 mM Bis-Tris Propane-HCl (pH 6.5 at 37°C),
10 mM MgCl₂,
0.1 mg/ml BSA B26 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

Buffer for Eco52I 10 mM Tris-HCl (pH 8.5 at 37°C),
3 mM MgCl₂,
100 mM NaCl,
0.1 mg/ml BSA B22 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for EcoRI 50 mM Tris-HCl (pH 7.5 at 37°C),
10 mM MgCl₂,
100 mM NaCl,
0.02% Triton X-100,
0.1 mg/ml BSA B12 5x1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for KpnI 10 mM Tris-HCl (pH 7.5 at 37°C),
10 mM MgCl₂,
0.02% Triton X-100,
0.1 mg/ml BSA B29 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for SdaI 37 mM Tris-acetate (pH 7.0 at 37°C),
15 mM magnesium acetate,
150 mM potassium acetate,
0.1 mg/ml BSA B24 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for SduI, Ppu21I 10 mM Tris-HCl (pH 7.2 at 37°C),
3 mM MgCl₂,
150 mM NaCl,
0.1 mg/ml BSA B23 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

10X Buffer for TaqI 10 mM Tris-HCl (pH 8.0 at 37°C),
5 mM MgCl₂,
100 mM NaCl,
0.1 mg/ml BSA B28 1 ml 10X buffer.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

Bovine Serum Albumin (BSA) 20 mg/ml solution in 10 mM Tris-HCl,
100 mM KCl,
1 mM EDTA,
50% glycerol (pH 7.4 at 25°C) B14 5mg.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

Dilution Buffer for Restriction Endonucleases 10 mM Tris-HCl,
100 mM KCl,
1 mM EDTA,
1 mM DTT,
50% glycerol,
200 µg/ml BSA (pH 7.4 at 25°C). B19 5x1 ml.
Certificate of Analysis
MSDS (English)
MSDS (English-USA)
MSDS (German)

Dilution of Restriction Enzymes

You may sometimes need to store diluted restriction enzymes for extended periods of time. For these purposes a single dilution/storage buffer is available for all Fermentas restriction enzymes:

10 mM Tris-HCl,

100 mM KCl,

1 mM EDTA,

1 mM DTT,

50% glycerol,

200 µg/ml BSA (pH 7.4 at 25°C).

Restriction enzymes with the same storage buffer composition (see product's Certificate of Analysis) as that of Dilution Buffer, retain their activity for at least 12 months when diluted with Dilution Buffer. All other restriction enzymes with different storage buffer requirements will retain more than 50% of their activity if stored at -20°C for 3-4 weeks after dilution with Dilution Buffer.

catalog@fermentas.com

Updated balandžio 25, 2008 08:51

Fermentas Buffer	1X Buffer Composition	Catalog #	Quantity Certificate of Analysis (in pdf, ~20 KB), MSDS
Buffer Set for Restriction Endonucleases		B30	1 ml 10X each of B, G, O, R, Tango™ buffers. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer B (blue)	10 mM Tris-HCl (pH 7.5 at 37°C), 10 mM MgCl₂, 0.1 mg/ml BSA	BB5	5x1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer G (green)	10 mM Tris-HCl (pH 7.5 at 37°C), 10 mM MgCl₂, 50 mM NaCl, 0.1 mg/ml BSA	BG5	5x1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer O (orange)	50 mM Tris-HCl (pH 7.5 at 37°C), 10 mM MgCl₂, 100 mM NaCl, 0.1 mg/ml BSA	BO5	5x1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer R (red)	10 mM Tris-HCl (pH 8.5 at 37°C), 10 mM MgCl₂, 100 mM KCl, 0.1 mg/ml BSA	BR5	5x1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer Tango™ (yellow)	33 mM Tris-acetate (pH 7.9 at 37°C), 10 mM magnesium acetate, 66 mM potassium acetate, 0.1 mg/ml BSA	BY5	5x1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for BamHI, Lsp1109I	10 mM Tris-HCl (pH 8.0 at 37°C), 5 mM MgCl₂, 100 mM KCl, 0.02% Triton X-100, 1 mM 2-mercapthoethanol, 0.1 mg/ml BSA	B57	5x1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for BfuI	50 mM Tris-acetate (pH 7.9 at 37°C), 15 mM Mg-acetate, 100 mM K-acetate, 0.1 mg/ml BSA	B59	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for AarI, AjiI, Bpu10I, ScaI, PasI	10 mM Bis-Tris Propane-HCl (pH 6.5 at 37°C), 10 mM MgCl₂, 100 mM KCl, 0.1 mg/ml BSA	B27	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for BseXI	50 mM Tris-HCl (pH 7.5 at 37°C), 2 mM MgCl₂, 100 mM NaCl, 0.1 mg/ml BSA	B31	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for Bsp143I	33 mM Tris-acetate (pH 7.9 at 37°C), 10 mM Mg-acetate, 66 mM K-acetate, 0.02% Triton X-100, 0.1 mg/ml BSA	B13	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for Cfr9I	10 mM Tris-HCl (pH 7.2 at 37°C), 5 mM MgCl₂, 200 mM sodium glutamate, 0.1 mg/ml BSA	B02	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for Cfr10I	10 mM Tris-HCl (pH 8.0 at 37°C), 5 mM MgCl₂, 100 mM NaCl, 0.02% Triton X-100, 0.1 mg/ml BSA	B04	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for Eam1105I	10 mM Tris-HCl (pH 7.5 at 37°C), 5 mM MgCl₂, 100 mM NaCl, 0.1 mg/ml BSA	B25	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for Ecl136II, SacI	10 mM Bis-Tris Propane-HCl (pH 6.5 at 37°C), 10 mM MgCl₂, 0.1 mg/ml BSA	B26	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
Buffer for Eco52I	10 mM Tris-HCl (pH 8.5 at 37°C), 3 mM MgCl₂, 100 mM NaCl, 0.1 mg/ml BSA	B22	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for EcoRI	50 mM Tris-HCl (pH 7.5 at 37°C), 10 mM MgCl₂, 100 mM NaCl, 0.02% Triton X-100, 0.1 mg/ml BSA	B12	5x1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for KpnI	10 mM Tris-HCl (pH 7.5 at 37°C), 10 mM MgCl₂, 0.02% Triton X-100, 0.1 mg/ml BSA	B29	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for SdaI	37 mM Tris-acetate (pH 7.0 at 37°C), 15 mM magnesium acetate, 150 mM potassium acetate, 0.1 mg/ml BSA	B24	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for SduI, Ppu21I	10 mM Tris-HCl (pH 7.2 at 37°C), 3 mM MgCl₂, 150 mM NaCl, 0.1 mg/ml BSA	B23	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
10X Buffer for TaqI	10 mM Tris-HCl (pH 8.0 at 37°C), 5 mM MgCl₂, 100 mM NaCl, 0.1 mg/ml BSA	B28	1 ml 10X buffer. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
Bovine Serum Albumin (BSA)	20 mg/ml solution in 10 mM Tris-HCl, 100 mM KCl, 1 mM EDTA, 50% glycerol (pH 7.4 at 25°C)	B14	5mg. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)
Dilution Buffer for Restriction Endonucleases	10 mM Tris-HCl, 100 mM KCl, 1 mM EDTA, 1 mM DTT, 50% glycerol, 200 µg/ml BSA (pH 7.4 at 25°C).	B19	5x1 ml. Certificate of Analysis MSDS (English) MSDS (English-USA) MSDS (German)