Thermo Scientific™ FastDigest XhoI

Thermo Scientific™ FastDigest XhoI is a high-performance, fast-acting restriction enzyme that recognizes and cleaves the palindromic sequence 5'- C↓TCGAG -3' / 3'- GAGC↑TC -5'. Designed for rapid and efficient DNA digestion within 5–15 minutes, this enzyme is fully compatible with the universal FastDigest and FastDigest Green buffers, enabling single, double, or multiple enzyme digestions without the need for buffer changes or additional DNA purification steps.

FastDigest XhoI ensures high specificity, no star activity, and 100% compatibility with downstream applications, making it ideal for molecular cloning, restriction mapping, genotyping, and other DNA modification protocols.

Key Features & Benefits

1. Ultra-Fast DNA Digestion in 5–15 Minutes
   • Accelerates molecular cloning workflows by reducing incubation time.
   • Ensures complete digestion in a single reaction step.
2. 100% Activity in Universal FastDigest Buffers
   • Eliminates the need for buffer changes or additional purification steps.
   • Works with FastDigest and FastDigest Green buffers, ensuring optimal performance.
3. No Star Activity for High-Specificity DNA Digestion
   • Designed to prevent nonspecific cleavage, maintaining precise restriction site integrity.
4. Direct Gel Loading with FastDigest Green Buffer
   • Allows immediate analysis without additional sample preparation.
   • Contains tracking dyes and density reagents for streamlined workflow.
5. Fully Compatible with DNA Modifying Enzymes
   • Supports direct addition of T4 DNA Ligase, Alkaline Phosphatase, Klenow Fragment, and T4 DNA Polymerase.
   • Ensures efficient DNA modification, ligation, and downstream processing.

Product Specifications

Use Cases & Applications

1. Molecular Cloning & Recombinant DNA Technology

• Plasmid Digestion for Vector Cloning
  • Efficiently cuts plasmid DNA for cloning and subcloning experiments.
  • Generates sticky ends for directional ligation of DNA inserts.
• High-Efficiency Double Digestion for DNA Fragment Cloning
  • Compatible with other FastDigest enzymes, reducing digestion time.
  • Supports seamless cloning workflows without buffer changes.

2. Restriction Site Mapping & Genetic Analysis

• Genotyping & SNP Detection
  • Used in mutation analysis and genetic variation studies.
  • Enables restriction fragment length polymorphism (RFLP) assays.
• Southern Blot Hybridization for DNA Fragment Analysis
  • Generates distinct restriction fragments for probe hybridization.
  • Ensures accurate identification of gene arrangements and mutations.

3. High-Throughput Cloning & Next-Generation Sequencing (NGS)

• Efficient Restriction Digestion for DNA Library Preparation
  • Supports NGS workflows by preparing DNA for sequencing adapters.
  • Ensures uniform fragment size distribution for quality sequencing data.
• Compatible with Automated Cloning Systems
  • Fast protocol eliminates extended incubation times.
  • Reduces sample handling and pipetting errors.

4. Epigenetics & Methylation-Sensitive Restriction Analysis

• CpG Methylation-Sensitive DNA Cleavage
  • Differentiates methylated vs. unmethylated DNA regions.
  • Useful for epigenetic regulation studies and DNA methylation profiling.

Recommended Protocol for FastDigest XhoI Digestion

1. Reaction Setup (50 μL Volume):
   • DNA Template: 0.5–1 μg
   • FastDigest XhoI (10 U/μL): 1 μL
   • 10X FastDigest Buffer: 5 μL
   • Nuclease-Free Water: Adjust to 50 μL
2. Incubation:
   • Digest at 37°C for 5–15 minutes.
3. Enzyme Inactivation (Optional):
   • Heat-inactivate at 80°C for 20 minutes, if needed.
4. Analysis of Digestion Products:
   • Load directly onto agarose gel (1-2%) using FastDigest Green Buffer.
   • Visualize using UV transillumination or fluorescence-based detection.

Storage & Handling Instructions

• Storage Conditions:
  • Store at -20°C for long-term enzyme stability.
  • Avoid freeze-thaw cycles to maintain enzyme activity.
• Handling Precautions:
  • Use nuclease-free water and buffers for optimal performance.
  • Work in a DNA-free environment to prevent contamination.
• Disposal Guidelines:
  • Dispose of used buffers and enzyme solutions according to biosafety waste regulations.

Why Choose Thermo Scientific™ FastDigest XhoI?

✅ Ultra-fast digestion (5–15 minutes) for rapid cloning workflows

✅ 100% activity in universal FastDigest Buffers—no buffer changes needed

✅ No star activity—ensures high-fidelity restriction site cleavage

✅ Direct gel loading with FastDigest Green Buffer—eliminates extra steps

✅ Compatible with downstream applications, including PCR, ligation, and sequencing

Thermo Scientific™ FastDigest XhoI is a high-efficiency restriction enzyme, enabling faster, more precise DNA digestion with reduced workflow complexity, making it ideal for cloning, sequencing, and molecular biology research.