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| Product Name | Fast Restriction Enzyme RsaI (Recognition Site: 5'-GT↓AC-3') |
| Catalog No. | IVDRM-0075 |
| Description | RsaI is a fast restriction endonuclease produced by recombinant genetic engineering. Working in a single universal buffer shared by the entire fast enzyme line, it delivers precise, complete DNA cleavage in just 5-15 minutes, greatly simplifying reaction setup and multi-enzyme digests. With generous activity headroom, it handles excess substrate and difficult templates with ease, making it ideal for rapid digestion of plasmid DNA, PCR products, and genomic DNA. RsaI specifically recognizes 5'-GT↓AC-3' and cleaves as indicated. |
| Intended Use | For rapid digestion of plasmid DNA, PCR products, and genomic DNA in molecular cloning and other molecular biology research applications. For research use only. |
| Principle / Technology | Restriction endonuclease expressed by recombinant genetic engineering; the enzyme specifically recognizes its palindromic/degenerate target sequence in double-stranded DNA and catalyzes phosphodiester bond cleavage at defined positions within the recognition site. |
| Sample Type | Plasmid DNA, PCR products, genomic DNA, and other double-stranded DNA substrates. |
| Performance Range / Specifications | Recognition site: 5'-GT↓AC-3' 3'-CA↑TG-5' Enzyme activity: 10 U/μL Recommended reaction conditions: 1× universal fast digestion buffer; incubate at 37°C. Inactivation: Cannot be heat inactivated; use phenol-chloroform extraction or column purification. Isoschizomers: AfaI, Csp6I, CviQI, RsaNI Number of recognition sites in common DNAs: λDNA 150, ΦX174 8, pBR322 12, pUC57 5, pUC18/19 5, SV40 11, M13mp18/19 4, Adeno2 78. Activity in different reaction buffers: universal fast digestion buffer 100%, commercial CutSmart-type buffer 100%, other commercial fast-digestion buffers ~50%. |
| Specificity | RsaI specifically recognizes and cleaves double-stranded DNA at: 5'-GT↓AC-3' 3'-CA↑TG-5' Isoschizomers: AfaI, Csp6I, CviQI, RsaNI |
| Reaction Conditions / Protocol | 1× universal fast digestion buffer; incubate at 37°C. Rapid DNA digestion (set up on ice): Plasmid DNA (20 μL total): ddH2O 15 μL, 10× universal fast digestion buffer (or color buffer) 2 μL, substrate DNA 2 μL (~1 μg), enzyme 1 μL. Incubate at 37°C for 15 min. PCR product (30 μL total): ddH2O 16 μL, 10× buffer 3 μL, substrate DNA 10 μL (~0.2 μg), enzyme 1 μL. Incubate at 37°C for 15-30 min. Genomic DNA (50 μL total): ddH2O 30 μL, 10× buffer 5 μL, substrate DNA 10 μL (5 μg), enzyme 5 μL. Incubate at 37°C for 30-60 min. *For unpurified PCR products (higher ionic strength), the buffer volume may be reduced to 2 μL. Purify PCR products before digestion if cloning follows. Mix gently (do not vortex) and briefly centrifuge. When the color buffer is used, digestion products can be loaded directly onto gels for electrophoresis. Note: This enzyme cannot be heat inactivated; terminate the reaction by phenol-chloroform extraction or column purification. |
| Storage Conditions | Store at -25 to -15°C. |
| Shelf Life | 2 years when stored at -25 to -15°C. |
| Package Specifications | 100 T |
| Concentration | 10 U/μL |
| Activity / Unit Definition | Enzyme activity: 10 U/μL. One unit is the amount of enzyme required for complete digestion of the substrate DNA under the standard fast-digestion reaction system. |
| Source / Origin | Recombinant expression by genetic engineering. |
| Expiration Date / Stability | Stable for 2 years at -25 to -15°C. |
| Compatibility | AfaI, Csp6I, CviQI, RsaNI |
| Recommended Buffer System | All fast restriction enzymes in this series share one universal digestion buffer, simplifying reaction setup and enabling convenient double or multiple digests. Relative activity: universal fast digestion buffer 100%, commercial CutSmart-type buffer 100%, other commercial fast-digestion buffers ~50%. |
| Application Notes / Precautions | Prepare reaction mixtures on ice. Mix gently by pipetting or flicking the tube; do not vortex. Briefly centrifuge to collect droplets. For double or multiple digests, use 1 μL of each enzyme and scale the reaction volume as needed; the total enzyme volume should not exceed 1/10 of the total reaction volume. If the enzymes used have different optimal reaction temperatures, start digestion with the enzyme requiring the lower temperature, then add the other enzyme(s) and incubate at the higher temperature. If the total reaction volume exceeds 20 μL, use a water bath, metal bath, or sand bath and extend the incubation time. |
| Recognition Site | 5'-GT↓AC-3' 3'-CA↑TG-5' |
| Heat Inactivation | Cannot be heat inactivated; use phenol-chloroform extraction or column purification to remove the enzyme. |
| Methylation Sensitivity | Dam, Dcm, EcoKI, EcoBI methylation: no effect. CpG methylation: cleavage affected. |
For research use only, not for clinical use.
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