Thermo Scientific™ AarI is a high-precision Type IIS restriction endonuclease designed for advanced molecular biology applications requiring cleavage outside the recognition sequence. AarI recognizes the asymmetric DNA sequence:

5′-CACCTGC(4/8)^-3′

3′-GTGGACG(8/4)^-5′

Unlike conventional Type II enzymes that cut within their recognition sites, AarI cleaves 4 nucleotides downstream on the top strand and 8 nucleotides downstream on the bottom strand, generating defined cohesive ends suitable for directional cloning strategies and engineered overhang assembly.

Supplied at 2 U/μL, AarI exhibits optimal activity at 37 °C in its dedicated 10X Buffer AarI, which contains premixed BSA for enhanced enzyme stability and protection against contaminants.

Mechanism & Performance Characteristics

Type IIS Restriction Behavior

AarI belongs to the Type IIS class of restriction enzymes. These enzymes:

• Recognize asymmetric DNA sequences
• Cleave outside their recognition sites
• Generate customizable overhangs
• Are ideal for modular cloning systems and precise DNA assembly

Because cleavage occurs outside the recognition sequence, the original recognition site can be removed from the final construct—an advantage in seamless cloning strategies.

Cleavage Efficiency Considerations

• Minimum two recognition sites required for efficient cleavage.
• For DNA containing a single AarI site, the addition of 0.5 μM oligonucleotide containing the AarI recognition sequence significantly improves digestion efficiency.
• Complete digestion of certain substrates may remain challenging due to enzyme–DNA interaction characteristics.
• Greater than 10-fold overdigestion may result in star activity (reduced specificity).

Gel Migration Note

AarI may remain associated with cleaved DNA fragments, potentially causing band shifting during electrophoresis.

Recommended Solution:

• Use 6X DNA Loading Dye & SDS Solution, or
• Heat digested samples in the presence of SDS before loading on agarose gels.

Applications

Thermo Scientific™ AarI is suitable for:

• Advanced molecular cloning
• Directional cloning workflows
• Restriction site mapping
• Genotyping
• Southern blotting
• Restriction Fragment Length Polymorphism (RFLP)
• SNP detection
• Custom overhang generation

Technical Specifications

Ordering Information

Storage Conditions: Store at –20 °C.

For Research Use Only. Not for diagnostic procedures.

Quality & Buffer System

Thermo Scientific restriction enzymes are manufactured under stringent quality control standards and are optimized for compatibility within the Five Buffer System. The inclusion of premixed BSA enhances enzyme stability and supports consistent digestion performance across diverse DNA preparations.

The universal Tango buffer is provided for convenience in double-digestion workflows, though AarI performs best in its dedicated buffer.

Thermo Scientific™ AarI (2 U/μL) is a reliable, high-specificity Type IIS restriction enzyme engineered for precise DNA manipulation. Its ability to cleave outside its recognition site makes it particularly valuable for advanced cloning strategies requiring defined overhangs and seamless construct assembly.

While AarI requires careful optimization—especially for single-site substrates—it delivers robust performance when used under recommended conditions. With heat inactivation capability, methylation sensitivity profiling, and compatibility within Thermo Scientific’s optimized buffer systems, AarI remains a powerful tool for molecular biologists engaged in complex DNA engineering workflows.