Product Catalog

Sequence specificity
Spe I recognizes the sequence *A↓*CTAGT and generates fragments with 5'-cohesive termini.

Source
From Sphaerotilus species

Compatible ends
Spe I ends are compatible with ends generated by Bln I, Nhe I, and Xba I.

Isoschizomers
The enzyme is an isoschizomer of Bcu I and Ahl I.

Methylation sensitivity
As indicated by (*) on the recognition sequence above, Spe I is inhibited by the presence of N6-methyladenine and 5’-methylcytosine ( ^mA↓^m CTAGT).

Activity in SuRE/Cut buffer System
Buffer printed in bold face type is the buffer recommended for optimal activity:

Incubation temperature
37°C

Unit definition
One unit is the enzyme activity that completely cleaves 1 µg Adenovirus 2 (Ad 2) DNA in 1 h at 37°C in a total volume of 25 µl SuRE/Cut buffer H.

Heat inactivation
Spe I (up to 100 U/µg DNA) can be heat-inactivated by incubating it for 15 min at 65°C.

Number of cleavage sites on different DNAs

1. Spe I
2. SuRE/Cut buffer H (10x)

Absence of nonspecific endonuclease activities
1 μg  Ad2 DNA  is incubated for 16 h in 50 μl SuRE/Cut buffer H with an excess of Spe I. The number of enzyme units that may be included in the incubation mixture without changing the enzyme-specific digestion pattern is stated under "Endo" (on the product label).

Absence of exonuclease activity
Approx. 5 μg [³H] labeled calf thymus DNA are incubated with 3 μl Spe I for 4 h at 37°C in a total volume of 100 μl 50 mM Tris-HCl, 10 mM MgCl₂, 1 mM dithioerythritol, pH approx. 7.5. The amount of radioactivity released is calculated as a percentage of input radioactivity, then divided by the number of units of enzyme. That calculated value is stated under "Exo" (on the product label).

Typical ligation and recutting assay
Spe I fragments obtained by complete digestion of 1 μg Ad2DNA are ligated for 16 h at 4°C with 1 U T4-DNA ligase  in 10 μl buffer that contains 66 mM Tris-HCl, 5 mM MgCl₂, 5 mM dithiothreitol, 1 mM ATP, pH 7.5 (at 20°C).
The percentages of product that can be ligated and subsequently recut with Spe I (yielding the typical pattern of Ad2 × Spe I fragments) are stated under "Lig" and "Rec" (on the product label).

• DNA Molecular Weight Marker XIV (100 bp ladder);
  
• Klenow Enzyme Labeling grade, from Escherichia coli lysogenic NM 964;
  
• Rapid DNA Ligation Kit;
  For rapid and efficient DNA ligation.