OPTIZYME™ AloI, Fisher BioReagents™

5'...^ ₇(N) G A A C (N)₆ T C C (N)_12-13^...3'
3'...^_12-13(N) C T T G (N)₆ A G G (N)₇ ^...5'

Supplied with: 10X OPTIZYME Buffer 5
Conditions for 100% Activity:

• 1X OPTIZYME Buffer 5: 10mM Tris-HCl (pH 8.5 at 37°C), 10mM MgCl₂, 100mM KCl and 0.1mg/mL BSA
• Incubate at 30°C

• Buffer 1: 0 - 20%
• Buffer 2: 0 - 20%
• Buffer 3: 0 - 20%
• Buffer 4: 20 - 50%
• Buffer 5: 100%

• 10mM Tris-HCl (pH 7.4 at 25°C), 100mM KCl, 1mM DTT, 1mM EDTA, 0.2mg/mL BSA and 50% (v/v) glycerol

• Incubation at 37°C results in 20% activity.
• AloI produces double-strand cuts on both sides from the interrupted recognition site. Its unique feature is a degenerate cleavage point on the 3 side of the recognition sequence (12 or 13 nucleotides away).
• The presence of S-adenosylmethionine in the reaction mixture results in incomplete cleavage with AloI.
• Greater than 10-fold overdigestion with AloI may result in star activity.
• AloI may remain associated with the cleaved DNA. This may cause DNA band shifting during electrophoresis. To avoid atypical DNA band patterns, use the 6X DNA Loading Dye & SDS Solution for sample preparation or heat the digested DNA in the presence of SDS prior to electrophoresis.

• 5'...GAAC(N)₄Gm6A TCC...3'

• 5'...GAAC(N)₆TCm5CW GG...3'

• 5'...m5C GAAC(N)₆TCC...3'

• 5'...GAAC(N)₆TCm5C G...3'

• 5'...GAAm5C G(N)₅TCC...3'