Xba I
ROCHE/XBAI-RO - from Xanthomonas campestris
Synonym: restriction enzyme
Product Type: Chemical
| biological source | Xanthomonas campestris |
| form | solution |
| manufacturer/tradename | Roche |
| packaging | pkg of 1,000 U (10674257001 [10 U/μl]) |
| pkg of 20,000 U (10674273001 [10 U/μl]) | |
| pkg of 20,000 U (11047663001 [40 U/μl]) | |
| pkg of 5,000 U (10674265001 [10 U/μl]) | |
| parameter | 37 °C optimum reaction temp. |
| Quality Level | 100  |
| storage temp. | −20°C |
| technique(s) | DNA sequencing: suitable |
| Analysis Note: | Absence of nonspecific endonuclease activities 1μg λDNA is incubated for 16 hours in 50 μl SuRE/Cut Buffer H with an excess of Xba 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 5 μg [3H] labeled calf thymus DNA are incubated with 3μl Xba I for 4 hours at +37°C in a total volume of 100 μl 50 mM Tris-HCl, 10 mM MgCl2, 1 mM Dithioerythritol, pH approximately 7.5. Under these conditions, no release of radioactivity is detectable, as stated in the certificate of analysis. |
| Analysis Note: | SuRE/Cut Buffer System The buffer in bold is recommended for optimal activity • A: 100% • B: 75-100% • H: 100% • L: 75-100% • M: 75-100% |
| Analysis Note: | Activity in PCR buffer: 60% Relative activity in PCR mix (Taq DNA Polymerase buffer) is 60%. The PCR mix contained λDNA, primers, 10 mM Tris-HCl (pH 8.3, 20 °C), 50 mM KCl, 1.5 mM MgCl2, 200 μM dNTPs, 2.5 U Taq DNA polymerase. The mix was subjected to 25 amplification cycles.Activity in reaction buffer of Pwo SuperYield DNA Polymerase PCR Mix is 25%. When supplemented with GC-RICH Solution, activity increases to 100%. Pwo SuperYield DNA Polymerase PCR Mix is not available in U.S. |
| Application: | The restriction enzyme Xba I has been used for the digestion of genomic DNA. |
| DNA Profile: | Number of cleavage sites on different DNAs • λ: 1 • φX174: 0 • Ad2: 5 • M13mp7: 0 • pBR322: 0 • pBR328: 0 • pUC18: 1 • SV40: 0 |
| General description: | Xba I recognizes the sequence T↓*CTAGA and generates fragments with 5′-cohesive termini. The enzyme needs at least two nucleotides around the target sequence before it will cut the cleavage site. Compatible ends Xba I ends are compatible with fragments generated by Avr I, Nhe I, and Spe I. Isoschizomers The enzyme is not known to have isoschizomers. Methylation sensitivity Xba I digestion of DNA is inhibited by the dam gene product of E. coli, which methylates the 6N position of adenine in the sequence GATC. The enzyme is also inhibited by 5-methylcytosine or 5-hydroxymethylcytosine at the site (*) indicated on the recognition sequence. Activity in SuRE/Cut Buffer System Buffer printed in bold face type is the buffer recommended for optimal activity: A B L M H 100% 75-100% 75-100% 75-100% 100% Relative activity in complete PCR mix Relative activity in PCR mix (Taq DNA Polymerase buffer) is 60%. The PCR mix contained ?DNA, primers, 10 mM Tris-HCl (pH 8.3, +20°C), 50 mM KCl, 1.5 mM MgCl2, 200 ?M dNTPs, 2.5 U Taq DNA polymerase. The mix was subjected to 25 amplification cycles. Activity in reaction buffer of Pwo SuperYield DNA Polymerase PCR Mix is 25%. When supplemented with GC-RICH Solution, activity increases to 100%. Incubation temperature +37°C Number of cleavage sites on different DNAs λ Ad2 SV40 ?X174 M13mp7 M13mp18 pBR322 pBR328 pUC18 1 5 0 0 0 1 0 0 1 PFGE tested Xba 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 10 U enzyme per ?g DNA and approximately 4 hour incubation time. Ligation and recutting assay Xba I fragments obtained by complete digestion of 1 ?g pUC18 DNA are ligated with 1 U T4 DNA Ligase in a volume of 10 ?l by incubation for 16 hours at +4°C in 66 mM Tris-HCl, 5 mM MgCl2, 5 mM Dithiothreitol, 1 mM ATP, pH 7.5 (at +20°C) resulting in >90% recovery of pUC18 DNA. Subsequent re-cutting with Xba I yields >95% of the typical pattern of pUC18 × Xba I fragments. |
| Other Notes: | For life science research only. Not for use in diagnostic procedures. |
| Specificity: | Star Activity The sequence specificity of Xba I is relaxed at low ionic strength or by addition of glycerol, ethanol or DMSO to the incubation mixture. Recognition sites: TCTAGA TCTAGA Restriction site: T↓CTAGA T↓CTAGA Heat inactivation: Xba I can be heat inactivated by incubation at 65 °C for 15 minutes (up to 15 U/μg DNA). Higher concentrations of Xba I cannot be heat inactivated completely under these conditions. |
| Unit Definition: | One unit is the enzyme activity that completely cleaves 1 μg λdam– DNA in one hour at +37 °C in a total volume of 50 μl (1x) 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 |