The GeneArt® Advanced Genetic Assembly System is a highly efficient kit for the simultaneous and seamless assembly of up to 10 DNA fragments (total length up to 110 Kbp) are assembled into any vector. The system relies on the yeast's ability to collect DNA fragments and recombine them with high efficiency. This greatly reduces in vitro processing of DNA, eliminating the need for enzymatic treatments (such as restriction enzyme digestion and ligation), while allowing precise fusion of DNA sequences. The kit contains materials for transformation and purification from yeast (including yeast selective media), and competent E. coli for plasmid amplification of correct cloning.

•Simple and effective—Clone up to 10 DNA fragments (containing the sequence of your choice) simultaneously in a single vector (up to 110 Kbp); no restriction enzyme digestion, Joining or Recombination Sites
•Precise and Efficient—Designed to enable you to clone the fragments you need, in the direction you need, at the site you need, and get up to 90% Correct cloning and no extra sequence left behind
•Flexibility—Use our linear vectors, a vector of your choice, or clone pre-existing end-to-end homologies without further modification DNA Fragments
•Free Tools—Use our free Web interfaceDesign DNA oligonucleotides and more
•More applications—Simplify a variety of synthetic biology and molecular biology techniques by quickly combining, adding, deleting, or exchanging DNA fragments

For cloning of 1 to 4 DNA fragments of limited size, and If you prefer the in vitro approach, consider using GeneArt® Seamless Cloning and assembly kit (Cat. No. A13288).

Easily create new specific constructs from multiple DNA fragments
GeneArt® high-order genetic assembly system utilizes yeast (Saccharomyces cerevisiae) Transformation-coupled recombination (TAR) ligates pre-existing DNA fragments or chemically synthesized oligonucleotides to a single recombinant molecule. Join DNA fragments and linearized vectors based on shared end homology. If such terminal homology does not exist between the two fragments, it can be "spliced" together with recombinant linkers, synthetic DNA oligonucleotides that provide terminal homology between two unrelated DNA fragments. The process is efficient and seamless, with no additional sequence after assembly. This product is optimized for use with up to 110 Kb and 10 DNA fragments in the vector, although it has been shown to support up to 0.5 Mb and 50 DNA fragments. situation.

Simple clone verification
To minimize yeast-specific work, recombinant yeast clones are extracted using a simplified 10-minute DNA extraction protocol. The assembled molecular weight is sufficient for colony PCR verification of junctions and direct transformation of E. coli cells for downstream analysis. Proprietary lysis buffer and glass beads required for extraction are included in the kit.

Considerations in Choosing Cloning Vectors
The GeneArt® High-Order Genetic Assembly System requires a shuttle with high capacity and compatible with yeast and E. coli Vector (i.e. BAC-YAC shuttle vector). This product does not include a cloning vector, but we provide a separate one called GeneArt® pYES1L Vector with Sapphire Technology™ (Cat. No. A13287). You can also use your own vector, but it must be compatible with higher-order genetic assembly systems. Compatibility is easily achieved with our GeneArt® Advanced Vector Transformation Cassette (Cat. No. A13291) .

Cloning Efficiency
In GeneArt® high-level assembly, the main factors affecting cloning efficiency are the size of the DNA element, the amount of DNA without terminal homology, and the total final Molecular size as well as fragment quality and specificity The ends of PCR fragments (A overhangs or blunt ends) do not affect cloning efficiency

Assembled into GeneArt® pYES1L Vector with Sapphire Technology™ with end homology. Typical cloning efficiencies for different number of fragments are as follows:
•For 5 DNA fragments (10 Kb each), >90%
• For 10 DNA fragments (5 Kb each), >90%
• For 10 DNA fragments (10 Kb each), >50%

assembled into GeneArt® pYES1L Vector with Sapphire Technology™ using 'spliced' DNA oligos without end homology Common cloning efficiencies for existing fragments are as follows:
• For 1 fragment (10 Kb), >90%
• For 2 DNA fragments (10 Kb each), >75%
< br>In silico clone design
GeneArt® A critical step in higher-order genetic assembly is the generation of fragments and oligonucleotides with appropriate homology and spacing Proper design to ensure successful assembly of clones. To simplify and speed up the design process, we offer aFree online toolsto help you design experiments through computer simulations. The tool checks experimental design for compatibility with product specifications, designs DNA oligonucleotides for PCR amplification or splicing of different elements for cloning, and shows the user a graphical representation of the vector and GenBank compatible with Vector NTI® software Downloadable annotation sequence in format.

Applications
The GeneArt® Advanced Genetic Assembly System is designed for cloning and DNA assembly experiments in a variety of molecular biology and synthetic biology applications. The product can be used to create modular expression vectors with interchangeable parts and can be used to perform a variety of tasks that might otherwise involve multiple steps. Use this kit to simplify: constructing fusion proteins; deleting, replacing, or adding DNA elements (such as restriction sites); cloning larger pre-existing DNA fragments that do not have terminal homology; and many other techniques that require manipulation of genetic sequences .