CloneSmart® Type IIS Assembly Kit – Cloning Kit
INTRODUCTION
CloneSmart® Type IIS Assembly Kit is a cloning tool based on the Golden Gate Assembly method, optimized for the combined use of the restriction enzyme BsaI and DNA ligase.
Because the recognition site of BsaI lies outside its cleavage site, it can generate customizable sticky ends (overhangs) — up to 256 possible types. When combined with DNA ligase, BsaI enables simultaneous cleavage and ligation reactions in a single tube (one‑step reaction).
During the reaction, the selective cleavage and continuous ligation mechanism within the thermal cycling program ensures that incorrectly ligated products are removed. This increases accuracy and reduces multimerization between DNA fragments. As a result, after approximately 1 hour of incubation, the reaction mixture is ready for transformation.
Principle of Operation:
- To assemble DNA fragments into a sequence or to insert one or multiple fragments into a vector, both the DNA inserts and plasmid must contain BsaI recognition sites, and the overhangs must be mutually compatible.
- For plasmids used in pre‑cloning or subcloning that lack BsaI sites, the kit can still be applied in combination with common Type IIP restriction enzymes (RE‑IIP) such as EcoRI, BamHI, SalI and HindIII.
- In such cases, DNA inserts are amplified using specially designed primers so that the 4‑bp overhangs generated by BsaI tạo ra tương thích với đầu 4 bp overhang do RE-IIP tạo ra, đồng thời một nucleotide được bổ sung trước trình tự 4 bp overhang trên mồi để ngăn chặn sự tái phân cắt tại các vị trí nhận biết RE-IIP.
CloneSmart® Type IIS Assembly Kit
HIGHLIGHTS
- High efficiency and accuracy: Assembly of single or multiple DNA fragments; customizable overhang design with high fidelity; scar‑free junctions.
- Broad enzyme compatibility and versatile applications: Works well with Type IIP restriction enzymes and is suitable for many common vector systems (pUC, pGEM, pET, pBAD).
- Simplified workflow and cost‑effective: Cleavage and ligation occur in the same reaction, reducing time and reagent consumption.
- Reduced multimerization: Eliminates undesired multimer formation between DNA fragments, improving target product yield and cloning efficiency.
SPECIFICATIONS
- Components: TypeIIS Enzyme Mix –Bsal, 10X TypeIIS Buffer, pGS Vector, Control Insert, Nuclease-free water
- Storage conditions: –20 °C
- Expiration date: 12 months from the date of manufacture
- Packaging: 10 tests/box
- Product code: PC‑02
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