Like our first-generation Seamless Cloning and Assembly Kits, the GeneArt™ Seamless PLUS Cloning and Assembly Kits are Complete kit for simultaneous directional cloning of 1 to 4 PCR fragments consisting of any sequence into any linearized vector in a single room temperature reaction of 30 minutes or less. However, the advantages of Seamless PLUS over previous kits are as follows:

•Improved efficiency:Pre-clones can be selected for large fragments to increase cloning efficiency
• < b>Larger Constructs:Create constructs up to 40 kb
• Versatility: High capacity for replication in most Gram-negative bacteria , a wide range of conjugation vectors

The above improvements are combined with these key benefits shared by all GeneArt™ Seamless Cloning and Assembly Kits:

•Quick and Easy — Clone up to 4 DNA fragments (containing the sequences of your choice) simultaneously in a single vector; no restriction enzymes, ligation or recombination sites required
• Precise and efficient — Designed to keep you Ability to clone the fragments you need, in the direction you need, at the site you need, and get up to 90% correct clones, with no extra sequence left behind
• Free Tools — Design your final construct and DNA in silico using our free web tool that walks you step-by-step through your plan Oligonucleotides
• Vector Flexibility — Use our linear vectors or your choice of vectors
• Multiple Applications — Quickly combine, add, Remove or swap DNA fragments to simplify many synthetic biology and molecular biology techniques

For cloning of more than 4 DNA fragments or where the final molecule is larger than 110 Kb, consider using GeneArt™ high-order genetic assembly system.

Easy and fast clone creation
GeneArt™ Seamless PLUS cloning is a simple two-step process that includes in vitro assembly and subsequent transformation into One Shot™ DH10B™ T1R SA Process for Competent E. coli. The kit uses a proprietary enzyme/buffer mixture to assemble DNA fragments with shared end homology, with no extra sequence or traces in the final construct ("seamless"). Terminal homology is easily incorporated through PCR amplification using custom DNA oligonucleotides (designed through our free web tools).

Efficient, flexible and precise cloning
When using the GeneArt™ Seamless PLUS Cloning and Assembly Kit, the main factor affecting cloning efficiency is DNA fragment size (100 bp to 10 Kb), total final molecular size (≤ 40 Kb), and the quality and specificity of each fragment.

Typical cloning efficiency for different number of fragments:

• For 4 fragments (5 Kb each), >95%
• For 4 fragments (10 Kb each), > ;90%

The success of cloning is independent of the insert sequence and vector type. You can design and add almost any desired sequence or sequence combination to any plasmid, as long as the plasmid can pass the restriction digestion Enzymatic digestion or PCR for linearization. The circular clone obtained from the reaction contains only your original vector sequence, insert, and specified homology, with no extraneous nucleotide insertions.

In silico Design Support
A critical step in GeneArt™ Seamless PLUS cloning is the correct design of fragments and oligonucleotides with appropriate homology and spacing, to help ensure successful cloning assembly. We offer a free online tool (GeneArt™ Design Tool for seamless or high-order assembly and mutation) to Helps you design experiments through computer simulations. The tool checks experimental design for compatibility with product specifications, designs DNA oligonucleotides with end homology for PCR amplification of different elements for cloning, and shows the user a graphical representation of the vector and its interaction with Vector NTI™ Downloadable annotated sequences in software-compatible GenBank format.

Applications
GeneArt™ Seamless PLUS Cloning and Assembly Kits are designed for cloning and DNA assembly in a variety of molecular and synthetic biology applications and beyond. . 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 kits to construct fusion proteins; delete, replace, or add DNA elements such as restriction sites in existing vectors; and perform a variety of other techniques that require manipulation of genetic sequences.