EcoR I

Isoschizomers: FunII

JBSpeed Restriction Enzyme

EcoRI

Catálogo Nº	Apresentação	Preço (R$)	Comprar
ENZ-105XS	5.000 U	262,50	Adicionar ao Carrinho
ENZ-105S	10.000 U	498,75	Adicionar ao Carrinho
ENZ-105M	20.000 U	949,20	Adicionar ao Carrinho
ENZ-105L	50.000 U	2.255,40	Adicionar ao Carrinho

5'–G↓AATT C–3'

3'–C TTAA↑G–5'

For in vitro use only!

Definição de unidade: One unit is the amount of enzyme required to completely digest 1 μg of Lambda DNA (5 sites) in 1 hour in a total reaction volume of 50 μl. Enzyme activity was determined in the recommended reaction buffer.

Envio: shipped on blue ice

Condições de armazenamento: store at -20 °C
avoid freeze/thaw cycles

Validade: 12 months

Forma: liquid (Supplied in 5 mM KPO₄ pH 7.4, 300 mM NaCl, 0.1 mM EDTA, 1 mM DTT, 200 μg/ml BSA, 0.15 % Triton X-100 and 50 % [v/v] glycerol)

Concentração: 10 units/μl

Source: Escherichia coli RY 13

Supplied with: 10x Universal Buffer (UB)

Recommended 50 μl assay

5 μl	10x Universal Buffer (UB)
1 μg	pure DNA¹ or PCR product²
10 units	enzyme
fill up to 50 μl	PCR grade water

¹ Supercoiled or high molecular weight DNA (e.g. plant genomic DNA) may require longer incubation time or higher amount of enzyme.
² Some enzymes may require additional DNA bases flanking the restriction site for complete digestion.

Protocol:

The enzyme should not exceed 10 % of total reaction volume.
Add enzyme as last component. Mix components well before adding enzyme. After enzyme addition, mix gently by pipetting. Do not vortex.
Incubate 5 to 10 min. at 37 °C.
Stop reaction by alternatively:
- Addition of 2.1 μl EDTA pH 8.0 [0.5 M], final 20 mM
- Heat Inactivation (20 min. at 65 °C)
- Spin Column DNA Purification (e.g. PCR Purification Kit, Cat.-No. PP-201S/L)
- Gel Electrophoresis and Single Band Excision (e.g. Agarose Gel Extraction Kit, Cat.-No. PP-202S/L)
- Phenol-Chloroform Extraction or Ethanol Precipitation.

Double Digestion - Buffer Compatibility:
B1 - 25-50 % Relative Activity
B2 - 50-75 % Relative Activity
B3 - 75 % Relative Activity
B4 - 50-75 % Relative Activity
B5 - 75 % Relative Activity
1x UB - 100 % Relative Activity (recommended)

Please note that the optimum digestion condition for this enzyme is 1x UB. Within the Universal Buffer (UB) system, the most majority of our enzymes display 100% Relative Activity in 1x UB and only few either in 0.5x or 2x UB. If optimum condition for second enzyme is different than the recommended for the first enzyme, we suggest carrying out first the restriction at the higher recommended concentration of UB and dilute the reaction volume to the adequate UB concentration for further proceeding with the second restriction.

Reaction Enzymes Buffer Guide:

Buffer 1	10 × B1	100 mM 100 mM 1000 μg/ml	Tris-HCl (pH 7.9, 25°C) MgCl₂ BSA
Buffer 2	10 × B2	100 mM 100 mM 500 mM 1000 μg/ml	Tris-HCl (pH 7.9, 25°C) MgCl₂ NaCl BSA
Buffer 3	10 × B3	500 mM 100 mM 1000 mM 1000 μg/ml	Tris-HCl (pH 7.9, 25°C) MgCl₂ NaCl BSA
Buffer 4	10 × B4	100 mM 100 mM 1500 mM 1000 μg/ml	Tris-HCl (pH 7.9, 25°C) MgCl₂ NaCl BSA
Buffer 5	10 × B5	200 mM 100 mM 500 mM 1000 μg/ml	Tris-acetate (pH 7.9, 25°C) Mg-acetate K-acetate BSA

Reaction Buffer Compatibility:
Our restriction enzymes are fully compatible to restrictases and buffer systems from other manufacturers and can be used along in double digestions. To obtain best results, consult the corresponding manuals of all involved products.

Ligation and recutting:
After 50-fold overdigestion with EcoRI, >98% of the DNA fragments can be ligated and recut with this enzyme.

Star activity:
Conditions of low ionic strength, high enzyme concentration, glycerol concentration >5 % or pH >8.0 may result in star activity.

DNA Methylation:
No Inhibition: dcm, dam
Inhibition (Impaired by overlapping): CpG

Quality Control:
All preparations are assayed for contaminating endonuclease, 3'-exonuclease, 5' exonuclease/ 5' phosphatase, as well as nonspecific single- and doublestranded DNase activities.

Produtos relacionados: Universal Restriction Buffer, #EN-300