Spe I
ROCHE/SPEI-RO - from Sphaerotilus species
Synonym: SPE I
Product Type: Chemical
| biological source | bacterial (Sphaerotilus spp.) |
| form | solution |
| manufacturer/tradename | Roche |
| packaging | pkg of 1,000 U (11008951001 [10 U/μl]) |
| pkg of 1,000 U (11207644001 [40 U/μl]) | |
| pkg of 200 U (11008943001 [10 U/μl]) | |
| parameter | 37 °C optimum reaction temp. |
| Quality Level | 100  |
| storage temp. | −20°C |
| Analysis Note: | 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). Incubation temperature +37°C PFGE tested Spe I has been tested in Pulsed-Field Gel Electrophoresis (on bacterial chromosomes). For cleavage of genomic DNA (E. coli C 600) embedded in agarose for PFGE analysis, we recommend using 10U of enzyme/μg DNA and 4 hour incubation time. Ligation and recutting assay Spe I fragments obtained by complete digestion of 1μg Ad2 DNA are ligated with 1U T4 DNA Ligase in a volume of 10μl by incubation for 16 hours at +4°C in 66mM Tris-HCl, 5mM MgCl2, 5mM Dithiothreitol, 1mM ATP, pH 7.5 (at +20°C), resulting in >90% recovery of Ad2 DNA. Subsequent re-cutting with Spe I yields >95% of the typical pattern of Ad2 × Spe I fragments. |
| Analysis Note: | SuRE/Cut Buffer System The buffer in bold is recommended for optimal activity • A: 75-100% • B: 75-100% • H: 100% • L: 75-100% • M: 100% |
| Analysis Note: | Activity in PCR buffer: Not tested |
| DNA Profile: | Number of cleavage sites on different DNAs • λ: 0 • φX174: 0 • Ad2: 3 • M13mp7: 0 • pBR322: 0 • pBR328: 0 • pUC18: 0 • SV40: 0 |
| General description: | Spe I recognizes the sequence *A↓*CTAGT and generates fragments with 5′-cohesive termini. |
| Other Notes: | For life science research only. Not for use in diagnostic procedures. |
| Quality: | Absence of nonspecific endonuclease activities 1μg Ad2 DNA is incubated for 16 hours in 50μl SuRE/Cut Buffer H with an excess of Spe I. The number of enzyme units which do not change the enzyme-specific pattern is stated in the certificate of analysis. Absence of exonuclease activity Approximately 5μg [3H] labeled calf thymus DNA are incubated with 3μl Spe I for 4 hours at +37°C in a total volume of 100μl 50mM Tris-HCl, 10mM MgCl2, 1mM Dithioerythritol, pH approximately 7.5. Under these conditions, no release of radioactivity is detectable, as stated in the certificate of analysis. |
| Specificity: | Recognition sites: *A*CTAGT *A*CTAGT Restriction site: *A↓*CTAGT *A↓*CTAGT Heat inactivation: Spe I can be heat inactivated by incubation at 65 °C for 15 minutes (up to 100 U/μg DNA). |
| Unit Definition: | One unit is the enzyme activity that completely cleaves 1 μg Ad2 DNA in one hour at +37 °C in a total volume of 25 μl SuRE/Cut Buffer H. |
| RIDADR | NONH for all modes of transport |
| WGK Germany | WGK 1 |
| Flash Point(F) | does not flash |
| Flash Point(C) | does not flash |
| Storage Temp. | −20°C |
| UNSPSC | 12352204 |
| Components | Enzyme Solution; SuRE/Cut Buffer H 10x concentrated |